CN1192647A

CN1192647A - Hypersensitive response induced resistance in plants

Info

Publication number: CN1192647A
Application number: CN96196146A
Authority: CN
Inventors: 魏钟民; S·V·比尔
Original assignee: Cornell Research Foundation Inc
Current assignee: Cornell Research Foundation Inc
Priority date: 1995-06-07
Filing date: 1996-06-05
Publication date: 1998-09-09
Anticipated expiration: 2016-06-05
Also published as: US5776889A; FI974430A; BR9609073A; AU714512B2; KR19990022577A; KR100529261B1; EP0871354A1; DK0871354T3; MX9709781A; DE69635307T2; WO1996039802A1; DE69635307D1; ES2248810T3; EP0871354B1; JPH11506938A; PL182459B1; FI974430A0; NZ309611A; CA2223616A1; US5859324A

Abstract

The present invention relates to a method of imparting pathogen resistance to plants. This involves applying a hypersensitive response elicitor polypeptide or protein in a non-infectious form to a plant under conditions where the polypeptide or protein contacts cells of the plant. The present invention is also directed to a pathogen resistant plant and a composition for imparting pathogen resistance to plants.

Description

The resistance that allergy is induced in plant corpus

The present invention finishes under the support of the competitive research foundation No.91-37303-6430 of the USDA NRI of U.S. government.

Invention field

The present invention relates to give the resistance that the plant allergy is induced.

Background of invention

The biochemical route that the organism evolution of living forms a series of complexity makes them discern and to reply the signal in the environment.These approach comprise the modification of acceptor organ, hormone, second messenger and enzyme.At present, about pathogene being attacked and made the signal transduction path that activates in the answering and know little about it, although this knowledge is most important to neurological susceptibility and the resistance of understanding disease plant.The general type of plant resistance to environment stress is that the restriction pathogene is in the internal breeding among a small circle around sites of infection.In many cases, this restriction is attended by local death's (that is necrosis) of host tissue.Integrate, pathogene restriction and local tissue necrosis have constituted anaphylactoid feature.Except local defense was replied, many plants were also replied by activating to make in the non-defence of infecting part of plant and to infection.As a result, whole strain plant has more resistance to secondary infection.This whole sustainable several weeks of resistance that obtain or longer time (R.E.F.Matternews phytovirology (academic press, New York, second edition, 1981)) and often give the cross tolerance to irrelevant pathogene (J.Kuc, plant disease control new method, I.Chet, compile (Wiley, New York, 1987), the 255-274 page or leaf is incorporated herein only for reference).

The failure of the tissue that the whole resistance expression that obtains and the invasion and attack of normal toxicity pathogene are immune relevant (" immunity that plant disease is induced ", bioscience, 32:854-856 (1982) quotes only for reference at this for Kuc, J.).The foundation of the whole resistance that obtains and the integral body of cell wall hydroxyprolin levels and peroxidase activity increase (Smith, J.A., Deng, " with cucumber; the comparative studies of the acid peroxidase that the resistance of inducing in muskmelon and the watermelon is relevant ", molecule physiology plant pathology, 14:329-338 (1988), quote only for reference at this) and the expression (Ward of the so-called whole resistant gene that obtains of 9 families of a cover, E.R., Deng, and " to induce the whole resistance reagent that obtains reply in the gene activity of collaborative (coordinate) ", plant cell, 3:49-59 (1991) is incorporated herein only for reference) interrelated.5 the still undetermined pathogenesis correlation of its physiological function of coding albumen in these defensin gene families.Yet; some has extracorporeal antifungal activity (Bol in these albumen; J.F.; Deng; " the plant pathogenesis correlation albumen of inducing by virus infections ", plant pathology yearbook, 28:113-38 (1990); be incorporated herein only for reference) and in transgene tobacco the constitutive expression of beans chitinase gene be protected from the infection (Broglie of fungi Rhizoctonia solani; K., etc., " having the genetically modified plants that fungal pathogens Rhizoctonia solani strengthened resistance "; science; 254:1194-1197 (1991) quotes only for reference at this), show that these whole resistance proteins that obtain can help immune state (Uknes; S.; Deng, " resistance of the acquisition in the Arabidopsis ", plant cell; 4:645-656 (1992) quotes only for reference at this).

As if salicylic acid play the effect of semiotic function because the endogenous level increases (Malamy after the immunity in inducing the whole resistance that obtains, J., Deng, " salicylic acid: a kind of possible endogenous signal in tobacco is replied the resistance of virus infections ", science 250:1002-1004 (1990), be incorporated herein only for reference) and exogenous salicylate induce the whole resistant gene (Yalpani that obtains, N., Deng, " salicylic acid is the inducer of pathogenesis associated protein in the tobacco of a kind of overall signal and virus infections ", plant cell 3:809-818 (1991) quotes only for reference at this), and obtain resistance (Uknes, S., etc., " the acquisition resistance in the Arabidopsis ", plant cell 4:645-656 (1992) is incorporated herein only for reference).In addition, salicylate wherein is encoded and does not demonstrate whole acquired resistance (Gaffney in the rotaring gene tobacco plant that the bacterium transgenosis of salicylate hydroxylase destroyed, T., Deng, " need salicylic acid to be used to induce globality to obtain resistance ", science 261:754-296 (1993) is incorporated herein only for reference).Yet, this effect may reflect the inhibition of part rather than overall signal function, and the detailed dynamic analysis of signal transmission shows that salicylate may be unnecessary (Rasmussen to the long range signals transmission in the cucumber, J.B., Deng, " integral body with salicylic acid accumulation in the cucumber of pathogenic mutation (the Pseudomonas Syringae pv.Syringae) inoculation of pseudomonas syringae back is induced ", plant physiology, 97:1342-1347) (1991) quote only for reference at this).

Broad research has been carried out in immunity with bioactivator.By in advance with cultivar-avirulence kind (Rahe, J.E., " resistance to beans anthracnose (Anthracnose) of in common kidney bean (Phaseolus Vulgaris), inducing ", plant pathology 59:1641-5 (1969); Elliston, J etc., " in the resistance to anthracnose of inducing ", plant pathology 61:1110-12 (1971) apart from a segment distance place that induces interaction sites; Skipp.R. etc., " research of cross protection in the beans anthracnose ", physiology plant pathology 3:299-313 (1973) quotes only for reference at this).Before occurring, symptom in host tissue, passes through heating and cultivar-pathogenicity kind (Rahe of perform toxic attenuation, J.E., Deng, " beans anthracnose thermal limit infects the metabolisming property of effect ", plant pathology: 60:1005-9 (1970), quote only for reference at this) or the non-pathogenic infection of beans, the disease that mung bean is caused with opposing Kidney bean hair disc spore (Colletotrichum lindemuthianum) cultivar cause of disease kind by immunity integrally.The Anthracnose Pathogen body of cucumber, Curcurbitaceae hair disc spore (Colletotrichumlagenarium) is effective equally as the inducer and the avirulence kind of anti-all kinds beans anthracnose Global Macros.Anti-one or more plant in cultivars of Kidney bean hair disc spore kinds and to the fungal species susceptible of all reports and therefore be called (Elliston in the cultivar of ' lacking genetic resistance ' pathogene, J., Deng; " beans avirulence hair disc spore belongs to the protection of kind to beans anthracnose ", Phytopathologische Zeitschrift 88:117-26 (1976); Elliston; J. etc.; " belong between kind and the Phaseolus Vulgaris compatible at the hair disc spore; the incompatible and comparative studies of inducing incompatible interaction to form "; PhytopathologischeZeitschrift 87:289-303 (1976) quotes only for reference at this) protection induced by Curcurbitaceae hair disc spore.These results show that identical mechanism can be induced (Elliston in the cultivar of " having " or " shortage " resistant gene of report; J; Deng, " relation of the accumulation of pisatin (Phytoalexin) and beans anti-anthracnose part and Global Macros " Phytopathologische Zeitschrift 88:114-30 (1977) is incorporated herein only for reference).It also is conspicuous that the cultivar of all kinds Kidney bean hair disc spore susceptible is had no lack of the gene that is used for the antipathogen resistance mechanism.

Kuc.J.; Deng; " by Curcurbitaceae hair disc spore protection cucumber opposing Curcurbitaceae hair disc spore "; physiology plant pathology 7:195-9 (1975); (quote at this only for reference) shows that cucumber plant can be by causing the disease that integrally protect its opposing to be caused by Curcurbitaceae hair disc spore with identical fungi inoculation cotyledon or rough leaf in advance.Therefore, cucumber is by integrally being protected its opposing fungi, bacterium and virus disease (Hammerschmidt with the local infection of fungi, bacterium or virus in advance, R. etc., " protection cucumber opposing Curcurbitaceae hair disc spore and cucumber branch spore (Cladosporium Cucumerinum) ", plant pathology 66:790-3 (1996); Jenns, A.E., etc., " with the local infection protection cucumber opposing Curcurbitaceae hair disc spore of tobacco necrosis virus ", physiology plant pathology 11:207-12 (1977); Caruso, F.L., etc., " cucumber of inducing by angular leaf spot of cucumber pseudomonad and Curcurbitaceae hair disc spore is to the resistance of anthracnose and angle tikka (Angular Leaf Spet) ", physiology plant pathology 14:191-201 (1979); Staub, T, etc., " by the disease that causes by cucumber branch spore and Curcurbitaceae hair disc spore with the opposing of the local infection Global Macros cucumber plant of arbitrary fungi in advance ", physiology plant pathology, 17:389-93 (1980); Bergstrom, G.C., etc., " by Curcurbitaceae hair disc spore, angular leaf spot of cucumber pseudomonad or tobacco necrosis virus local infection cucumber are to the influence of the whole resistance of cucumber mosaic virus ", plant pathology 72:922-6 (1982); Gessler, C., etc., " in cucumber, inducing the withered resistance of Fusarium ", plant pathology 72:1439-41 (1982) by the pathogene of root and leaf; Basham, B, etc., " tobacco necrosis virus is induced the whole resistance to Siberian cocklebur monofilament shell bacterium in cucumber ", physiology plant pathology 23:137-44 (1983) quotes only for reference at this).Infect the non-specific protection of being induced by Curcurbitaceae hair disc spore or tobacco necrosis virus and resist at least 13 kinds of pathogene effectively, comprise single-minded and the facultative parasitism fungi, local damage and whole virus, withered fungi (wilt fungi) and bacterium.Similarly, the root pathogene is induced and also resisted effectively to protection by the root pathogene.Other cucurbitaceous plant comprises that watermelon and muskmelon integrally protect its opposing Curcurbitaceae hair disc spore (Caruso, F.L.; Deng; " by Curcurbitaceae hair disc spore protection watermelon and muskmelon opposing Curcurbitaceae hair disc spore ", plant pathology 67:1285-9 (1977) quotes only for reference at this).

Global Macros in tobacco is also by the multiple disease of reactance (Kuc.J., etc., " immunity is used for the resistance of disease in tobacco ", tobacco science latest developments 9:179-213 (1983) are incorporated herein only for reference).The gangrenosum acne damage that causes by tobacco mosaic virus strengthened the disease that upper blade causes this virus resistance (Ross, A.F., etc., " resistance that in plant, obtains ", virology 14:340-58 (1961) by the infection induced integral body of local virus; Ross, A.F., Deng, " the globality influence that local damage forms ",: plant virus 127-50 page or leaf (1966), be incorporated herein only for reference), it is the pathogenic mutation (Phytophthoraparasitica var.nicotianae) of phytophthora parasitica, aphid (aphid) Myzus persicae (McIntyre of P.tabacina and wildfire pseudomonad (Pseudomonas tabaci) and reduction breeding, J.L., Deng, " by part and Global Macros " to the tobacco mosaic virus infection reactance phytophthora parasitica (Phytophthora Parasitica var.nicotianae) of the irritated tobacco of virus, physiology plant pathology 15:321-30 (1979); McIntyre, J.L., etc., " tobacco mosaic virus is to the influence to the whole resistance of resisting various pathogene and insect of the local infection of tobacco (Nicotiana Tabacum) ", plant pathology 71:297-301 (1981) quotes only for reference at this).Heat-killed P.tabaci (Lovrekovich, L., Deng, " in tobacco leaf, induce the reaction of anti-wildfire " with heat-killed bacterial treatment, nature 205:823-4 (1965), quote only for reference at this), and aeruginosa eggplant (Sequeira, L. etc., " interaction of bacterium and host cell wall: the relation between itself and the induction of resistance mechanism ", physiology plant pathology 10:43-50 (1977) quotes only for reference at this), immerse the resistance of inducing in the tobacco leaf to the identical bacterium that is used to immerse.Tobacco plant is also protected its opposing phytophthora parasitica (Pparasitica var.nicotiana) (McIntyre by nematode Pratylenchus penetrans; J.L.; Deng; " by cultivar-avirulence kind; acellular ultrasonic degradation thing and Pratylenchus Penetrans protection tobacco opposing phytophthora parasitica (Phytophthora Parasitica Var.Nicotianae) "; plant pathology 68:235-9 (1978) quotes only for reference at this).

Cruikshank, I.A.M., Deng, " stem of tobacco is invaded for the influence of blade to blue mould reaction " with Peronospora Tabacina Adam, the magazine 26:369-72 of agriculture in Australia Science Institute (1960) is incorporated herein only for referencely, is that first report is by with the anti-blue mould of fungi stem injecting immune tobacco leaf (promptly, P.tabacina), it also relates to dwarfing and old and feeble in advance.Recent findings has not only alleviated developmental arrest but also has accelerated g and D from the outside to the injection of xylem.The tobacco plant that immunity is crossed; no matter in the still open-air experiment in greenhouse; increase approximately 40%, dry weight increases by 40%, and fresh weight increases by 30%; and than many 4-6 of adjoining tree sheet leaf (Tuzun; S, etc., " with Peronospora Tabacina stem injection and Metalaxyl processing to the growth of tobacco in the field and protect it to resist the influence of blue mould "; plant pathology 74:804 (1984) is incorporated herein only for reference).These plant blossom comparisons are according to the about 2-3 week (Tuzun early of plant; S., etc., " activity of the factor in the tobacco that its Peronospora Tabacina Adam who resists blue mould with Global Macros infects "; physiology plant pathology 26:321-30 (1985) quotes only for reference at this).

Global Macros is not given the absolute immunity of infecting, but is reduced to severity of disease and postpones the formation of symptom.Damage number, lesion size, and the degree of fungal pathogens sporulation all reduce.Diseased region can reduce above 90%.

When imposing " reinforcement " inoculation the 3-6 week of cucumber behind primary vaccination; the immunity of being induced by Curcurbitaceae hair disc spore lasts till blooms and result (Kuc.J.; Deng; " by the aspect of Curcurbitaceae hair disc spore protection cucumber opposing Curcurbitaceae hair disc spore "; plant pathology 67:533-6 (1977) quotes only for reference at this).In case beginning to form fruit, plant can not induce protection.Tobacco plant the season of growth by with in addition immunity of the sporangium stem of P.tabacina injection.Yet,, be higher than 20cm effectively just this technology is only worked as plant in order to prevent the formation of whole blue mould.

From immune cucumber plant, remove the immune level induce leaf to have the stretching, extension leaf before not reducing.Yet later on the blade that occurs from terminal bud is than its other previous blade become gradually not protected (Dean, R.A.; Deng; " Global Macros of in cucumber, inducing: the time of generation and ' signal ' move ", plant pathology 76:966-70 (1986) quotes only for reference at this).Ross, A.F has reported similar result, and " group effect that local damage forms ": plant virus 127-50 page or leaf (1966), be incorporated herein only for reference, the immunity tobacco (local damage host) by resisting tobacco mosaic virus with tobacco mosaic virus infection in advance.On the contrary; the young leaves that the sprout that downcuts from the tobacco plant of stem's immunity with the P.tabacina injection sends is to be subjected to the highly (Tuzun of protection; S.; Deng; " by induce in the callus tobacco to blue mould (Peronospora Tabacina Adam; the transfer of resistance ", plant pathology 75:1304 (1985) is incorporated herein only for reference).The plant that regenerates by tissue culture from the leaf of immune plant is showing significant minimizing with respect to the plant from the maternal leaf regeneration of non-immunity aspect the blue mould.Young regenerate only shows the sporulation of minimizing.Along with plant is aging, damage forms and sporulation all reduces.Yet; other researcher does not lead to the same conclusion; although in an experiment, reported the remarkable minimizing (Lucas of sporulation; J.A.; Deng; " to the induction of resistance of Peronospora Hyoscyami from the not transferability of protected tobacco explant " to regenerate, plant pathology 75:1222-5 (1985) quotes only for reference at this).

The protection of cucumber and watermelon in greenhouse and field be effectively (Caruso, F.L., etc., " by the open-air protection of Curcurbitaceae hair disc spore cucumber opposing Curcurbitaceae hair disc spore ", plant pathology 67:1290-2 (1977) quotes only for reference at this).In single test, total damaged area of Curcurbitaceae hair disc spore is less than not protecting 2% of check plant damaged area on the protection cucumber.Similarly, 66 strains protection, only 1 strain death in the plant under fire, opposite 69 strains are not protected, 47 strain death in the watermelon under fire.In Kentucky and Puerto Rico widely in the field test, tobacco stem is injected in the blue mould of control at least and best fungicide with the sporangium of P.tabacina, and metalaxyl is effective equally.According to necrosis area and sporulation degree, plant 95-99% is protected, and the output that causes curing tobacco 10-25% increases.

As if the induction of resistance of antibacterium and virus show as inhibition that disease symptoms or pathogene duplicate or both (Caruso that haves both at the same time, F.L., Deng, " by angular leaf spot of cucumber pseudomonas and Curcurbitaceae hair disc spore inducing cucumber resistance " to anthracnose and angle tikka, physiology plant pathology 14:191-201 (1979); Doss, M., etc., " the whole resistance to the angular leaf spot of cucumber pseudomonas that obtains of cucumber shows as the inhibition of symptom, rather than the duplicating of bacterium ", AcatPhytopathologia Academiae Sclentiarum Hungaricae16:(3-4), 269-72 (1981); Jenns, A.E. etc., " by the non-specific resistance to pathogene of inducing " with tobacco necrosis virus, Curcurbitaceae hair disc spore or angular leaf spot of cucumber pseudomonad local infection cucumber integral body, Phytopathologia Mediterranea 18:129-34 (1979) quotes only for reference at this).

As the above, the research that obtains resistance about integral body comprises with infectious pathogen infection plant.Although the research in this field is that what how to be worked is useful for understanding the whole resistance that obtains, with infectious activating agent induce such resistance commercial be otiose because the contact of such phytopathogen can weaken or kill plants.The present invention relates to overcome this kind defective.

Summary of the invention

The present invention relates to give the method for plant pathogen resistance.The method is included in hypersensitive response elicitor (the hypersensitive response elicitor) polypeptide of using non-infectious form under the condition of polypeptide or albumen contact plant cell or albumen in plant.

Another aspect of the present invention relates to the pathogen-resistance plant, and its cell contacts with non-infectious hypersensitive response elicitor polypeptide or albumen.

Another aspect of the present invention relates to the composition that is used to give plant pathogen resistance.That said composition comprises is non-infectious, polypeptide or albumen and carrier are induced in allergy.

The present invention has following ability: the plant disease that treatment can not be treated in the past; Wholistic therapy is not because the high people of expense think the disease of treatment separately; And the infectious activating agent of avoiding using the treatment disease.The present invention can give resistance and need not to use to being had pathogenic activating agent by the plant treated or near be positioned at these treatment plants plant.Owing to the present invention includes the natural prodcuts of using fully biodegradable, so will not pollute the environment.

The accompanying drawing summary

Fig. 1 has shown that coding separates starch Erwinia hypersensitive response elicitor polypeptide or the albumen (that is hrpN) genetic structure of gene cluster.Top line shows the Restriction Enzyme collection of illustrative plates of plasmid vector pCPP 430, E=EcoRI wherein, B=BamHI, and H=HindIII.Rectangle is represented transcript unit, and the arrow below the rectangle is represented transcriptional orientation.Bigger arrow represents to be used for hypersensitive response elicitor polypeptide or the necessary zone of the final translation of albumen.PCPP 430 hrpN are derivatives of pCPP430, and hrpN is wherein suddenlyd change by inserting transposons TnStac.

Fig. 2 is the collection of illustrative plates of plasmid vector pCPP 9.Notable attribute is mobilization (mob) site that is used to connect; λ (cos) cos site; And be used for the separated region (par) of plasmid genetic stability.B, BamHI; E, EcoRI; H, HindIII; P, PstI; S, SaII; Sm, SmaI; OriV, origin of replication; Sp ^r, the spectinomycin resistance; Sm ^r, streptomycin resistance.

Detailed Description Of The Invention

The present invention relates to give the method for plant pathogen resistance. The method is included in polypeptide or albumen Under the condition of contact plant all or part of cell with a kind of hypersensitive response elicitor of non-infectious form Polypeptide or albumen are applied on all or part of plant.

Another aspect of the present invention relates to the hypersensitive response elicitor polypeptide of its cell and non-infectious form Or the pathogen-resistance plant of albumen contact.

A further aspect of the invention relates to be used to the composition of giving plant pathogen resistance. This group Compound comprises hypersensitive response elicitor polypeptide or albumen and the carrier of non-infection.

Being used for hypersensitive response elicitor polypeptide of the present invention or albumen can be corresponding to being derived from multiple pathogens Hypersensitive response elicitor polypeptide or albumen. This peptide species or albumen can planting at the contact inducer Induce local necrosis in the fabric texture. Preferred pathogen comprises separates starch Erwinia (Erwinia Amylovora), chrysanthemum Erwinia (Erwinia chrysanthemi), pseudomonas syringae (Pseudomonas syringae), aeruginosa eggplant (Pseudomonas solancearum), field Wild Xanthomonas campestris (Xanthomonas campestris), or its mixture.

For purpose of the present invention, but the hypersensitive response elicitor polypeptide of non-infectious form or albumen induced hypersensitivity reaction and not causing contacts the disease in the plant of this polypeptide or albumen.This can finish by many methods, comprising: 1) use inducer polypeptide or the albumen that separates; 2) use and not cause disease and with the bacterium of the genetic transformation of coding hypersensitive response elicitor polypeptide or albumen; With 3) be applied in and cause disease and the natural bacterium that contains coding hypersensitive response elicitor polypeptide or protein gene in some floristics (but not being those kinds that will use this bacterium).

In a specific embodiments of the present invention, the separable certainly corresponding organism of hypersensitive response elicitor polypeptide or protein also is applied to plant.This separating method is well-known, as described in the following document: Arlat, M., F.Van Gijsegem, J.C.Huet, J.C.Pemollet, and C.A.Boucher, " PopA1, a kind of albumen of induced hypersensitivity sample reaction in special Petunia genotype is by the Hrp approach secretion of aeruginosa eggplant ", EMBO is (1994) J.13:543-553; He, S.Y., H.C.Huang, and A.collmer, " Pseudomonassyringae pv.Syringae Harpin _Pss: a kind of albumen by secretion of Hrp approach and induced hypersensitivity reaction in plant ", cell 73:1255-1266 (1993); And Wei, Z.-M., R.J.Laby, C.H.Zumoff, D.W.Bauer, S.-Y.He, A.Collmer, and S.V.Beer, " separate the allergy Harpin inducer that the starch Erwinia produces by phytopathogen ", science 257:85-88 (1992) quotes only for reference at this.Also referring to unsettled U.S. Patent application series Nos.08/200,024 and 08/062,024, quote only for reference at this.Yet preferably, hypersensitive response elicitor polypeptide that the present invention separates or albumen are through the reorganization preparation and by following purifying.

In other embodiments of the present invention, hypersensitive response elicitor polypeptide of the present invention or albumen can be applied to plant by the bacterium that application contains the gene of coding hypersensitive response elicitor polypeptide or albumen.This bacterium must secrete or export polypeptide or thereby albumen makes inducer can contact plant cell.In these embodiments, hypersensitive response elicitor polypeptide or albumen in plant corpus (inplanta) or just before bacterium imports plant, produce by this bacterium.

In an embodiment of bacterium application model of the present invention, this bacterium does not cause disease and with the genetic transformation (for example, reorganization) of coding hypersensitive response elicitor polypeptide or albumen.For example, the Escherichia coli of induced hypersensitivity reaction not in plant, the genetic transformation of anaphylactoid inducer polypeptide of available code or albumen and be applied to plant then.Bacterial species (non-Escherichia coli) also can be used in this embodiment of the present invention.

In another embodiment of bacterium application model of the present invention, this bacterium does not cause disease and the natural gene that contains coding hypersensitive response elicitor polypeptide or albumen.The example of this bacterium explanation in front.Yet these bacteriums are used for the disease of this spread of germs plant of susceptible not in this embodiment.For example, separating the starch Erwinia causes disease but do not cause disease in tomato in apple or pears.Yet this bacterium will induced hypersensitivity reaction in tomato.Therefore, according to this embodiment of the present invention, separate the starch Erwinia and can be used for tomato and in this kind, do not cause disease to give pathogen-resistance.

Hypersensitive response elicitor polypeptide or albumen from the chrysanthemum Erwinia have the following amino acid sequence corresponding to Sequence Identification No.1: Met Gln Ile Thr Ile Lys Ala His Ile Gly Gly Asp Leu Gly Val Ser1 5 10 15Gly Leu Gly Ala Gln Gly Leu Lys Gly Leu Asn Ser Ala Ala Ser Ser

20 25 30Leu?Gly?Ser?Ser?Val?Asp?Lys?Leu?Ser?Ser?Thr?Ile?Asp?Lys?Leu?Thr

35 40 45Ser?Ala?Leu?Thr?Ser?Met?Met?Phe?Gly?Gly?Ala?Leu?Ala?Gln?Gly?Leu

50 55 60Gly?Ala?Ser?Ser?Lys?Gly?Leu?Gly?Met?Ser?Asn?Gln?Leu?Gly?Gln?Ser65 70 75 80Phe?Gly?Asn?Gly?Ala?Gln?Gly?Ala?Ser?Asn?Leu?Leu?Ser?Val?Pro?Lys

85 90 95Ser?Gly?Gly?Asp?Ala?Leu?Ser?Lys?Met?Phe?Asp?Lys?Ala?Leu?Asp?Asp

100 105 110Leu?Leu?Gly?His?Asp?Thr?Val?Thr?Lys?Leu?Thr?Asn?Gln?Ser?Asn?Gln

115 120 125Leu?Ala?Asn?Ser?Met?Leu?Asn?Ala?Ser?Gln?Met?Thr?Gln?Gly?Asn?Met

130 135 140Asn?Ala?Phe?Gly?Ser?Gly?Val?Asn?Asn?Ala?Leu?Ser?Ser?Ile?Leu?Gly145 150 155 160Asn?Gly?Leu?Gly?Gln?Ser?Met?Ser?Gly?Phe?Ser?Gln?Pro?Ser?Leu?Gly

165 170 175Ala?Gly?Gly?Leu?Gln?Gly?Leu?Ser?Gly?Ala?Gly?Ala?Phe?Asn?Gln?Leu

180 185 190Gly?Asn?Ala?Ile?Gly?Met?Gly?Val?Gly?Gln?Asn?Ala?Ala?Leu?Ser?Ala

195 200 205Leu?Ser?Asn?Val?Ser?Thr?His?Val?Asp?Gly?Asn?Asn?Arg?His?Phe?Val

210 215 220Asp?Lys?Glu?Asp?Arg?Gly?Met?Ala?Lys?Glu?Ile?Gly?Gln?Phe?Met?Asp225 230 235 240Gln?Tyr?Pro?Glu?Ile?Phe?Gly?Lys?Pro?Glu?Tyr?Gln?Lys?Asp?Gly?Trp

245 250 255Ser?Ser?Pro?Lys?Thr?Asp?Asp?Lys?Ser?Trp?Ala?Lys?Ala?Leu?Ser?Lys

260 265 270Pro?Asp?Asp?Asp?Gly?Met?Thr?Gly?Ala?Ser?Met?Asp?Lys?Phe?Arg?Gln

275 280 285Ala?Met?Gly?Met?Ile?Lys?Ser?Ala?Val?Ala?Gly?Asp?Thr?Gly?Asn?Thr

290 295 300Asn?Leu?Asn?Leu?Arg?Gly?Ala?Gly?Gly?Ala?Ser?Leu?Gly?Ile?Asp?Ala305 310 315 320Ala?Val?Val?Gly?Asp?Lys?Ile?Ala?Asn?Met?Ser?Leu?Gly?Lys?Leu?Ala

This hypersensitive response elicitor polypeptide of 325 330 335Asn Ala or albumen have the molecular weight of 34kDa; Are heat-staple; have the glycine content greater than 16%, and are substantially free of cysteine.No.2DNA：CGATTTTACC CGGGTGAACG TGCTATGACC GACAGCATCA CGGTATTCGA CACCGTTACG 60GCGTTTATGG CCGCGATGAA CCGGCATCAG GCGGCGCGCT GGTCGCCGCA ATCCGGCGTC 120GATCTGGTAT TTCAGTTTGG GGACACCGGG CGTGAACTCA TGATGCAGAT TCAGCCGGGG 180CAGCAATATC CCGGCATGTT GCGCACGCTG CTCGCTCGTC GTTATCAGCA GGCGGCAGAG 240TGCGATGGCT GCCATCTGTG CCTGAACGGC AGCGATGTAT TGATCCTCTG GTGGCCGCTG 300CCGTCGGATC CCGGCAGTTA TCCGCAGGTG ATCGAACGTT TGTTTGAACT GGCGGGAATG 360ACGTTGCCGT CGCTATCCAT AGCACCGACG GCGCGTCCGC AGACAGGGAA CGGACGCGCC 420CGATCATTAA GATAAAGGCG GCTTTTTTTA TTGCAAA CG GTAACGGTGA GGAACCGTTT 480CACCGTCGGC GTCACTCAGT AACAAGTATC CATCATGATG CCTACATCGG GATCGGCGTG 540GGCATCCGTT GCAGATACTT TTGCGAACAC CTGACATGAA TGAGGAAACG AAATTATGCA 600AATTACGATC AAAGCGCACA TCGGCGGTGA TTTGGGCGTC TCCGGTCTGG GGCTGGGTGC 660TCAGGGACTG AAAGGACTGA ATTCCGCGGC TTCATCGCTG GGTTCCAGCG TGGATAAACT 720GAGCAGCACC ATCGATAAGT TGACCTCCGC GCTGACTTCG ATGATGTTTG GCGGCGCGCT 780GGCGCAGGGG CTGGGCGCCA GCTCGAAGGG GCTGGGGATG AGCAATCAAC TGGGCCAGTC 840TTTCGGCAAT GGCGCGCAGG GTGCGAGCAA CCTGCTATCC GTACCGAAAT CCGGCGGCGA 900TGCGTTGTCA AAAATGTTTG ATAAAGCGCT GGACGATCTG CTGGGTCATG ACACCGTGAC 960CAAGCTGACT AACCAGAGCA ACCAACTGGC TAATTCAATG CTGAACGCCA GCCAGATGAC 1020CCAGGGTAAT ATGAATGCGT TCGGCAGCGG TGTGAACAAC GCACTGTCGT CCATTCTCGG 1080CAACGGTCTC GGCCAGTCGA TGAGTGGCTT CTCTCAGCCT TCTCTGGGGG CAGGCGGCTT 1140GCAGGGCCTG AGCGGCGCGG GTGCATTCAA CCAGTTGGGT AATGCCATCG GCATGGGCGT 1200GGGGCAGAAT GCTGCGCTGA GTGCGTTGAG TAACGTCAGC ACCCACGTAG ACGGTAACAA 1260CCGCCACTTT GTAGATAAAG AAGATCGCGG CATGGCGAAA GAGATCGGCC AGTTTATGGA 1320TCAGTATCCG GAAATATTCG GTAAACCGGA ATACCAGAAA GATGGCTGGA GTTCGCCGAA 1380GACGGACGAC AAATCCTGGG CTAAAGCGCT GAGTAAACCG GATGATGACG GTATGACCGG 1440CGCCAGCATG GACAAATTCC GTCAGGCGAT GGGTATGATC AAAAGCGCGG TGGCGGGTGA 1500TACCGGCAAT ACCAACCTGA ACCTGCGTGG CGCGGGCGGT GCATCGCTGG GTATCGATGC 1560GGCTGTCGTC GGCGATAAAA TAGCCAACAT GTCGCTGGGT AAGCTGGCCA ACGCCTGATA 1620ATCTGTGCTG GCCTGATAAA GCGGAAACGA AAAAAGAGAC GGGGAAGCCT GTCTCTTTTC 1680TTATTATGCG GTTTATGCGG TTACCTGGAC CGGTTAATCA TCGTCATCGA TCTGGTACAA 1740ACGCACATTT TCCCGTTCAT TCGCGTCGTT ACGCGCCACA ATCGCGATGG CATCTTCCTC 1800GTCGCTCAGA TTGCGCGGCT GATGGGGAAC GCCGGGTGGA ATATAGAGAA ACTCGCCGGC 1860CAGATGGAGA CACGTCTGCG ATAAATCTGT GCCGTAACGT GTTTCTATCC GCCCCTTTAG 1920CAGATAGATT GCGGTTTCGT AATCAACATG GTAATGCGGT TCCGCCTGTG CGCCGGCCGG 1980GATCACCACA ATATTCATAG AAAGCTGTCT TGCACCTACC GTATCGCGGG AGATACCGAC 2040AAAATAGGGC AGTTTTTGCG TGGTATCCGT GGGGTGTTCC GGCCTGACAA TCTTGAGTTG 2100GTTCGTCATC ATCTTTCTCC ATCTGGGCGA CCTGATCGGT T 2141

Hypersensitive response elicitor polypeptide or the albumen of explaining the starch Erwinia by oneself have the following amino acid sequence corresponding to Sequence Identification No.3: Met Ser Leu Asn Thr Ser Gly Leu Gly Ala Ser Thr Met Gln Ile Ser1 5 10 15Ile Gly Gly Ala Gly Gly Asn Asn Gly Leu Leu Gly Thr Ser Arg Gln

20 25 30Asn?Ala?Gly?Leu?Gly?Gly?Asn?Ser?Ala?Leu?Gly?Leu?Gly?Gly?Gly?Asn

35 40 45Gln?Asn?Asp?Thr?Val?Asn?Gln?Leu?A?la?Gly?Leu?Leu?Thr?Gly?Met?Met

50 55 60Met?Met?Met?Ser?Met?Met?Gly?Gly?Gly?Gly?Leu?Met?Gly?Gly?Gly?Leu65 70 75 80Gly?Gly?Gly?Leu?Gly?Asn?Gly?Leu?Gly?Gly?Ser?Gly?Gly?Leu?Gly?Glu

85 90 95Gly?Leu?Ser?Asn?Ala?Leu?Asn?Asp?Met?Leu?Gly?Gly?Ser?Leu?Asn?Thr

100 105 110Leu?Gly?Ser?Lys?Gly?Gly?Asn?Asn?Thr?Thr?Ser?Thr?Thr?Asn?Ser?Pro

115 120 125Leu?Asp?Gln?Ala?Leu?Gly?Ile?Asn?Ser?Thr?Ser?Gln?Asn?Asp?Asp?Ser

130 135 140Thr?Ser?Gly?Thr?Asp?Ser?Thr?Ser?Asp?Ser?Ser?Asp?Pro?Met?Gln?Gln145 150 155 160Leu?Leu?Lys?Met?Phe?Ser?Glu?Ile?Met?Gln?Ser?Leu?Phe?Gly?Asp?Gly

165 170 175Gln?Asp?Gly?Thr?Gln?Gly?Ser?Ser?Ser?Gly?Gly?Lys?Gln?Pro?Thr?Glu

180 185 190Gly?Glu?Gln?Asn?Ala?Tyr?Lys?Lys?Gly?Val?Thr?Asp?Ala?Leu?Ser?Gly

195 200 205Leu?Met?Gly?Asn?Gly?Leu?Ser?Gln?Leu?Leu?Gly?Asn?Gly?Gly?Leu?Gly

210 215 220Gly?Gly?Gln?Gly?Gly?Asn?Ala?Gly?Thr?Gly?Leu?Asp?Gly?Ser?Ser?Leu225 230 235 240Gly?Gly?Lys?Gly?Leu?Gln?Asn?Leu?Ser?Gly?Pro?Val?Asp?Tyr?Gln?Gln

245 250 255Leu?Gly?Asn?Ala?Val?Gly?Thr?Gly?Ile?Gly?Met?Lys?Ala?Gly?Ile?Gln

260 265 270Ala?Leu?Asn?Asp?Ile?Gly?Thr?His?Arg?His?Ser?Ser?Thr?Arg?Ser?Phe

275 280 285Val?Asn?Lys?Gly?Asp?Arg?Ala?Met?Ala?Lys?Glu?Ile?Gly?Gln?Phe?Met

290 295 300Asp?Gln?Tyr?Pro?Glu?Val?Phe?Gly?Lys?Pro?Gln?Tyr?Gln?Lys?Gly?Pro305 310 315 320Gly?Gln?Glu?Val?Lys?Thr?Asp?Asp?Lys?Ser?Trp?Ala?Lys?Ala?Leu?Ser

325 330 335Lys?Pro?Asp?Asp?Asp?Gly?Met?Thr?Pro?Ala?Ser?Met?Glu?Gln?Phe?Asn

340 345 350Lys?Ala?Lys?Gly?Met?Ile?Lys?Arg?Pro?Met?Ala?Gly?Asp?Thr?Gly?Asn

355 360 365Gly?Asn?Leu?Gln?His?Ala?Val?Pro?Val?Val?Leu?Arg?Trp?Val?Leu?Met

This hypersensitive response elicitor polypeptide of 370 375 380Pro385 or albumen have the molecular weight of about 37kDa; And its pI is about 4.3; 100 ℃ of of of of of of at least of and its, 10 minutes be heat-staple.This hypersensitive response elicitor polypeptide or albumen do not have cysteine basically.Explain by oneself in the hypersensitive response elicitor polypeptide of starch Erwinia or the albumen document below and described in more detail; Be i.e. Wei; Z.-m.; R.J.Laby; C.H.ZumofF; D.W.Bauer; S.-Y.He; A.Collmer; And S.V.Beer; " Harpin separates the hypersensitive response elicitor that the starch Erwinia produces by phytopathogen ", science 257:85-88 ( 1992 ) quotes only for reference at this.DNA4：ATGAGTCTGA ATACAAGTGG GCTGGGAGCG TCAACGATGC AAATTTCTAT CGGCGGTGCG 60GGCGGAAATA ACGGGTTGCT GGGTACCAGT CGCCAGAATG CTGGGTTGGG TGGCAATTCT 120GCACTGGGGC TGGGCGGCGG TAATCAAAAT GATACCGTCA ATCAGCTGGC TGGCTTACTC 180ACCGGCATGA TGATGATGAT GAGCATGATG GGCGGTGGTG GGCTGATGGG CGGTGGCTTA 240GGCGGTGGCT TAGGTAATGG CTTGGGTGGC TCAGGTGGCC TGGGCGAAGG ACTGTCGAAC 300GCGCTGAACG ATATGTTAGG CGGTTCGCTG AACACGCTGG GCTCGAAAGG CGGCAACAAT 360ACCACTTCAA CAACAAATTC CCCGCTGGAC CAGGCGCTGG GTATTAACTC AACGTCCCAA 420AACGACGATT CCACCTCCGG CACAGATTCC ACCTCAGACT CCAGCGACCC GATGCAGCAG 480CTGCTGAAGA TGTTCAGCGA GATAATGCAA AGCCTGTTTG GTGATGGGCA AGATGGCACC 540CAGGGCAGTT CCTCTGGGGG CAAGCAGCCG ACCGAAGGCG AGCAGAACGC CTATAAAAAA 600GGAGTCACTG ATGCGCTGTC GGGCCTGATG GGTAATGGTC TGAGCCAGCT CCTTGGCAAC 660GGGGGACTGG GAGGTGGTCA GGGCGGTAAT GCTGGCACGG GTCTTGACGG TTCGTCGCTG 720GGCGGCAAAG GGCTGCAAAA CCTGAGCGGG CCGGTGGACT ACCAGCAGTT AGGTAACGCC 780GTGGGTACCG GTATCGGTAT GAAAGCGGGC ATTCAGGCGC TGAATGATAT CGGTACGCAC 840AGGCACAGTT CAACCCGTTC TTTCGTCAAT AAAGGCGATC GGGCGATGGC GAAGGAAATC 900GGTCAGTTCA TGGACCAGTA TCCTGAGGTG TTTGGCAAGC CGCAGTACCA GAAAGGCCCG 960GGTCAGGAGG TGAAAACCGA TGACAAATCA TGGGCAAAAG CACTGAGCAA GCCAGATGAC 1020GACGGAATGA CACCAGCCAG TATGGAGCAG TTCAACAAAG CCAAGGGCAT GATCAAAAGG 1080CCCATGGCGG GTGATACCGG CAACGGCAAC CTGCAGCACG CGGTGCCGGT GGTTCTTCGC 1140TGGGTATTGA TGCCATGA 1158

Has following amino acid sequence from the hypersensitive response elicitor polypeptide of pseudomonas syringae or albumen: Met Gln Ser Leu Ser Leu Asn Ser Ser Ser Leu Gln Thr Pro Ala Met1 5 10 15Ala Leu Val Leu Val Arg Pro Glu Ala Glu Thr Thr Gly Ser Thr Ser corresponding to Sequence Identification numbers 5

20 25 30Ser?Lys?Ala?Leu?Gln?Glu?Val?Val?Val?Lys?Leu?Ala?Glu?Glu?Leu?Met

35 40 45Arg?Asn?Gly?Gln?Leu?Asp?Asp?Ser?Ser?Pro?Leu?Gly?Lys?Leu?Leu?Ala

50 55 60?Lys?Ser?Met?Ala?Ala?Asp?Gly?Lys?Ala?Gly?Gly?Gly?Ile?Glu?Asp?Val65 70 75 80Ile?Ala?Ala?Leu?Asp?Lys?Leu?Ile?His?Glu?Lys?Leu?Gly?Asp?Asn?Phe

85 90 95Gly?Ala?Ser?Ala?Asp?Ser?Ala?Ser?Gly?Thr?Gly?Gln?Gln?Asp?Leu?Met

100 105 110Thr?Gln?Val?Leu?Asn?Gly?Leu?Ala?Lys?Ser?Met?Leu?Asp?Asp?Leu?Leu

115 120 125Thr?Lys?Gln?Asp?Gly?Gly?Thr?Ser?Phe?Ser?Glu?Asp?Asp?Met?Pro?Met

130 135 140Leu?Asn?Lys?Ile?Ala?Gln?Phe?Met?Asp?Asp?Asn?Pro?Ala?Gln?Phe?Pro145 150 155 160Lys?Pro?Asp?Ser?Gly?Ser?Trp?Val?Asn?Glu?Leu?Lys?Glu?Asp?Asn?Phe

165 170 175Leu?Asp?Gly?Asp?Glu?Thr?Ala?Ala?Phe?Arg?Ser?Ala?Leu?Asp?Ile?Ile

180 185 190Gly?Gln?Gln?Leu?Gly?Asn?Gln?Gln?Ser?Asp?Ala?Gly?Ser?Leu?Ala?Gly

195 200 205Thr?Gly?Gly?Gly?Leu?Gly?Thr?Pro?Ser?Ser?Phe?Ser?Asn?Asn?Ser?Ser

210 215 220Val?Met?Gly?Asp?Pro?Leu?Ile?Asp?Ala?Asn?Thr?Gly?Pro?Gly?Asp?Ser225 230 235 240Gly?Asn?Thr?Arg?Gly?Glu?Ala?Gly?Gln?Leu?Ile?Gly?Glu?Leu?Ile?Asp

245 250 255Arg?Gly?Leu?Gln?Ser?Val?Leu?Ala?Gly?Gly?Gly?Leu?Gly?Thr?Pro?Val

260 265 270Asn?Thr?Pro?Gln?Thr?gly?Thr?Ser?Ala?Asn?Gly?Gly?Gln?Ser?Ala?Gln

275 280 285Asp?Leu?Asp?Gln?Leu?Leu?Gly?Gly?Leu?Leu?Leu?Lys?Gly?Leu?Glu?Ala

290 295 300Thr?Leu?Lys?Asp?Ala?Gly?Gln?Thr?Gly?Thr?Asp?Val?Gln?Ser?Ser?Ala305 310 315 320Ala?Gln?Ile?Ala?Thr?Leu?Leu?Val?Ser?Thr?Leu?Leu?Gln?Gly?Thr?Arg

325 330 335Asn?Gln?Ala?Ala?Ala

340 these hypersensitive response elicitor polypeptide or albumen have the molecular weight of 34-35kDa.It is rich in glycine (about 13.5%) and lacks cysteine and tyrosine.Relevant further INFORMATION DISCOVERY from the pseudomonas syringae hypersensitive response elicitor is in He, S.Y., H.C.Huang, and A.Collmer, " Pseudomonas syringae pv.Syringae Harpin _Pss: a kind of albumen by secretion of Hrp approach and induced hypersensitivity reaction in plant ", cell 73:1255-1266 ( 1993 ) quotes only for reference at this.DNA6：ATGCAGAGTC TCAGTCTTAA CAGCAGCTCG CTGCAAACCC CGGCAATGGC CCTTGTCCTG 60GTACGTCCTG AAGCCGAGAC GACTGGCAGT ACGTCGAGCA AGGCGCTTCA GGAAGTTGTC 120GTGAAGCTGG CCGAGGAACT GATGCGCAAT GGTCAACTCG ACGACAGCTC GCCATTGGGA 180AAACTGTTGG CCAAGTCGAT GGCCGCAGAT GGCAAGGCGG GCGGCGGTAT TGAGGATGTC 240ATCGCTGCGC TGGACAAGCT GATCCATGAA AAGCTCGGTG ACAACTTCGG CGCGTCTGCG 300GACAGCGCCT CGGGTACCGG ACAGCAGGAC CTGATGACTC AGGTGCTCAA TGGCCTGGCC 360AAGTCGATGC TCGATGATCT TCTGACCAAG CAGGATGGCG GGACAAGCTT CTCCGAAGAC 420GATATGCCGA TGCTGAACAA GATCGCGCAG TTCATGGATG ACAATCCCGC ACAGTTTCCC 480AAGCCGGACT CGGGCTCCTG GGTGAACGAA CTCAAGGAAG ACAACTTCCT TGATGGCGAC 540GAAACGGCTG CGTTCCGTTC GGCACTCGAC ATCATTGGCC AGCAACTGGG TAATCAGCAG 600AGTGACGCTG GCAGTCTGGC AGGGACGGGT GGAGGTCTGG GCACTCCGAG CAGTTTTTCC 660AACAACTCGT CCGTGATGGG TGATCCGCTG ATCGACGCCA ATACCGGTCC CGGTGACAGC 720GGCAATACCC GTGGTGAAGC GGGGCAACTG ATCGGCGAGC TTATCGACCG TGGCCTGCAA 780TCGGTATTGG CCGGTGGTGG ACTGGGCACA CCCGTAAACA CCCCGCAGAC CGGTACGTCG 840GCGAATGGCG GACAGTCCGC TCAGGATCTT GATCAGTTGC TGGGCGGCTT GCTGCTCAAG 900GGCCTGGAGG CAACGCTCAA GGATGCCGGG CAAACAGGCA CCGACGTGCA GTCGAGCGCT 960GCGCAAATCG CCACCTTGCT GGTCAGTACG CTGCTGCAAG GCACCCGCAA TCAGGCTGCA 1020

Has following amino acid sequence from the hypersensitive response elicitor polypeptide of eggplant pseudomonas or albumen: Met Ser Val Gly Asn Ile Gln Ser Pro Ser Asn Leu Pro Gly Leu Gln1 5 10 15Asn Leu Asn Leu Asn Thr Asn Thr Asn Ser Gln Gln Ser Gly Gln Ser corresponding to Sequence Identification numbers 7

20 25 30Val?Gln?Asp?Leu?Ile?Lys?Gln?Val?Glu?Lys?Asp?Ile?Leu?Asn?Ile?Ile

35 40 45Ala?Ala?Leu?Val?Gln?Lys?Ala?Ala?Gln?Ser?Ala?Gly?Gly?Asn?Thr?Gly

50 55 60Asn?Thr?Gly?Asn?Ala?Pro?Ala?Lys?Asp?Gly?Asn?Ala?Asn?Ala?Gly?Ala65 70 75 80Asn?Asp?Pro?Ser?Lys?Asn?Asp?Pro?Ser?Lys?Ser?Gln?Ala?Pro?Gln?Ser

85 90 95Ala?Asn?Lys?Thr?Gly?Asn?Val?Asp?Asp?Ala?Asn?Asn?Gln?Asp?Pro?Met

100 105 110Gln?Ala?Leu?Met?Gln?Leu?Leu?Glu?Asp?Leu?Val?Lys?Leu?Leu?Lys?Ala

115 120 125Ala?Leu?His?Met?Gln?Gln?Pro?Gly?Gly?Asn?Asp?Lys?Gly?Asn?Gly?Val

130 135 140Gly?Gly?Ala?Asn?Gly?Ala?Lys?Gly?Ala?Gly?Gly?Gln?Gly?Gly?Leu?Ala145 150 155 160Glu?Ala?Leu?Gln?Glu?Ile?Glu?Gln?Ile?Leu?Ala?Gln?Leu?Gly?Gly?Gly

165 170 175Gly?Ala?Gly?Ala?Gly?Gly?Ala?Gly?Gly?Gly?Val?Gly?Gly?Ala?Gly?Gly

180 185 190Ala?Asp?Gly?Gly?Ser?Gly?Ala?Gly?Gly?Ala?Gly?Gly?Ala?Asn?Gly?Ala

195 200 205Asp?Gly?Gly?Asn?Gly?Val?Asn?Gly?Asn?Gln?Ala?Asn?Gly?Pro?Gln?Asn

210 215 220Ala?Gly?Asp?Val?Asn?Gly?Ala?Asn?Gly?Ala?Asp?Asp?Gly?Ser?Glu?Asp225 230 235 240Gln?Gly?Gly?Leu?Thr?Gly?Val?Leu?Gln?Lys?Leu?Met?Lys?Ile?Leu?Asn

245 250 255Ala?Leu?Val?Gln?Met?Met?Gln?Gln?Gly?Gly?Leu?Gly?Gly?Gly?Asn?Gln

260 265 270Ala?Gln?Gly?Gly?Ser?Lys?Gly?Ala?Gly?Asn?Ala?Ser?Pro?Ala?Ser?Gly

275 280 285Ala?Asn?Pro?Gly?Ala?Asn?Gln?Pro?Gly?Ser?Ala?Asp?Asp?Gln?Ser?Ser

290 295 300Gly?Gln?Asn?Asn?Leu?Gln?Ser?Gln?Ile?Met?Asp?Val?Val?Lys?Glu?Val305 310 315 320Val?Gln?Ile?Leu?Gln?Gln?Met?Leu?Ala?Ala?Gln?Asn?Gly?Gly?Ser?Gln

325 330 335Gln?Ser?Thr?Ser?Thr?Gln?Pro?Met

340 its by following have corresponding to the dna molecular of the nucleotide sequence of Sequence Identification numbers 8 coded: ATGTCAGTCG GAAACATCCA GAGCCCGTCG AACCTCCCGG GTCTGCAGAA CCTGAACCTC 60AACACCAACA CCAACAGCCA GCAATCGGGC CAGTCCGTGC AAGACCTGAT CAAGCAGGTC 120GAGAAGGACA TCCTCAACAT CATCGCAGCC CTCGTGCAGAAG GCCGCACA GTCGGCGGGC 180GGCAACACCG GTAACACCGG CAACGCGCCG GCGAAGGACG GCAATGCCAA CGCGGGCGCC 240AACGACCCGA GCAAGAACGA CCCGAGCAAG AGCCAGGCTC CGCAGTCGGC CAACAAGACC 300GGCAACGTCG ACGACGCCAA CAACCAGGAT CCGATGCAAG CGCTGATGCA GCTGCTGGAA 360GACCTGGTGA AGCTGCTGAA GGCGGCCCTG CACATGCAGC AGCCCGGCGG CAATGACAAG 420GGCAACGGCG TGGGCGGTGC CAACGGCGCC AAGGGTGCCG GCGGCCAGGG CGGCCTGGCC 480GAAGCGCTGC AGGAGATCGA GCAGATCCTC GCCCAGCTCG GCGGCGGCGG TGCTGGCGCC 540GGCGGCGCGG GTGGCGGTGT CGGCGGTGCT GGTGGCGCGG ATGGCGGCTC CGGTGCGGGT 600GGCGCAGGCG GTGCGAACGG CGCCGACGGC GGCAATGGCG TGAACGGCAA CCAGGCGAAC 660GGCCCGCAGA ACGCAGGCGA TGTCAACGGT GCCAACGGCG CGGATGACGG CAGCGAAGAC 720CAGGGCGGCC TCACCGGCGT GCTGCAAAAG CTGATGAAGA TCCTGAACGC GCTGGTGCAG 780ATGATGCAGC AAGGCGGCCT CGGCGGCGGC AACCAGGCGC AGGGCGGCTC GAAGGGTGCC 840GGCAACGCCT CGCCGGCTTC CGGCGCGAAC CCGGGCGCGA ACCAGCCCGG TTCGGCGGAT 900GATCAATCGT CCGGCCAGAA CAATCTGCAA TCCCAGATCA TGGATGTGGT GAAGGAGGTC 960GTCCAGATCC TGCAGCAGAT GCTGGCGGCG CAGAACGGCG GCAGCCAGCA GTCCACCTCG 1020ACGCAGCCGA TGTAA 1035 is about setting forth in the further information document below from aeruginosa eggplant hypersensitive response elicitor polypeptide or albumen: Arlat, M., F.Van Gijsegem, J.C.Huet, J.C.Pemollet, and C.A.Boucher, " PopA1; a kind of albumen of induced hypersensitivity sample reaction in special Petunia genotype; " by the Hrp approach secretion of aeruginosa eggplant, EMBD magazine .13:543-533 (1994) quotes only for reference at this.

Has following amino acid sequence from the cause a disease hypersensitive response elicitor polypeptide of mutation glycine or albumen of Xanthomonas campestris: Thr Leu Ile Glu Leu Met Ile Val Val Ala Ile Ile Ala Ile Leu Ala1 5 10 15Ala Ile Ala Leu Pro Ala Tyr Gln Asp Tyr corresponding to Sequence Identification numbers 9

20 25 these sequences are amino terminal sequences, only have 26 residues from pathogenic mutation glycine hypersensitive response elicitor polypeptide of Xanthomonas campestris or albumen.It is complementary with the pili protein subunit of measuring in other Xanthomonas campestris causes a disease mutation.

Top inducer is exemplary.Other inducer can be identified by the bacterium of cultivating the induced hypersensitivity reaction under the situation about expressing in the coding stimulant gene.Acellular goods in the culture supernatant can detect its inducer activity (that is local necrosis) by immersing suitable plant tissue with them.

It also is possible that the fragment of top hypersensitive response elicitor polypeptide or albumen and other pathogene total length inducer fragment are used for method of the present invention.

Suitable fragment can prepare by several method.In first method, the subclone of the known inducer protein gene of encoding is by the conventional molecular genetic operation preparation of subclone genetic fragment.This subclone can be tested the more small protein or the peptide of inducer activity at external or bacterial cell expression in vivo according to following method to produce then.

As selection, the inducer protein fragments can be by preparing with proteolytic enzyme such as chymotrypsin or SP A or tryptic digestion total length inducer albumen.Different proteolytic enzymes may cut inducer albumen in different sites according to the amino acid sequence of inducer albumen.Some fragment that proteolysis obtains can be the activity inducement thing of resistance.

In another kind of scheme, according to the primary structure knowledge of albumen, the fragment of inducer protein gene can be synthesized together by the concrete primer cover with round pcr and this albumen specific part of representative of selecting.These fragments will be cloned into peptide or the albumen that appropriate carriers is used to increase and express brachymemma then.

Variant also can pass through (or as selecting), and for example, disappearance or interpolation are modified the amino acid that polypeptide characteristic, secondary structure and hydrotherapy character have minimum influence.For example, polypeptide can be connected on signal (or leading) sequence at the N-of albumen end, and this sequence is translated or anti-translation altogether, the transhipment of mediation albumen.Polypeptide also can be connected on attachment or other sequence so that synthetic, the purifying of polypeptide or evaluation.

Albumen of the present invention or polypeptide preferably by routine techniques with the form of purifying (preferably at least about 80%, more preferably 90% purity) preparation.Usually, the secretion of albumen of the present invention or polypeptide enters the growth medium of recombination bacillus coli.In order to separate this albumen, breeding has e. coli host cell, the homogenate of recombinant plasmid, and centrifugal homogenate product is to remove bacterial debris.Then supernatant is carried out ammonium sulfate precipitation subsequently.The part that will contain polypeptide of the present invention or albumen is carried out gel filtration with protein isolate in the glucan in suitable aperture or polyacrylamide post.If necessary, protein part can be further purified by HPLC.

The dna molecular of coding hypersensitive response elicitor polypeptide or albumen can mix in the cell with the recombinant DNA technology of routine.In general, this comprises that it is in heterologous (that is, not being normal presence) the expression integral body to it that dna molecular is inserted this dna molecular.This allogeneic dna sequence DNA molecule inserts with suitable significant direction and correct reading frame and expresses integral body.This carrier contains the albumen coded sequence that is useful on insertion and transcribes and translate necessary element.

The U.S. Patent No. 4,237,224 of Cohen and Boyer is quoted only for referencely at this, described with Restriction Enzyme cutting and the preparation with the expression integral body of recombinant plasmid form that is connected with dna ligase.These recombinant plasmids import by method for transformation then and duplicate in comprising the prokaryotes that grow in the tissue culture and eukaryotic unicellular culture.

Recombination gene also can import virus, as vaccinia virus.Recombinant virus can produce by plasmid transfection being advanced in the cell with virus infections.

Suitable carriers includes, but are not limited to following viral vectors such as the whole gt11 of λ carrier, gtWES.tB, Charon 4 and plasmid vector such as pBR 322, pBR 325, pACYC 177, and pACYC 184, and pUC 8, pUC 9, and pUC 18, and pUC 19, pLG 339, and pR 290, and pKC 37, pKC 101, and SV 40, pBluescript II SK+/-or KS+/-(referring to " clone's integral body of Stratagene " catalogue (1993), Stratagene, La Jolla, Calif quotes only for reference at this), pQE, pIH 821, pGEX, and pET series is (referring to F.WStudier etc., " with the expression of T7 RNA polymerase mediation clone gene ", gene expression technique 185 is rolled up (1990), quotes only for reference at this), and any derivative.Recombinant molecule can be by transforming, especially transduce, engage, moving or the electroporation transfered cell.Dna sequence dna is cloned into carrier with the cloning process of this area standard, as described in Maniatis etc., and molecular cloning: experiment guide, cold spring harbor laboratory, the cold spring port, New York (1982) quote only for reference at this.

Many host's carrier integral body can be utilized with the expressing protein coded sequence.Mainly, carrier integral body must be compatible with used host cell.Host-vector integral body includes but not limited to as follows: with the bacterium of phage DNA, plasmid DNA or cosmid DNA conversion; Microorganism such as the yeast that contains yeast vector; Mammalian cell integral body with virus (for example: vaccinia virus, adenovirus etc.) infection; The insect cell integral body that infects with virus (as, baculoviral); Reach plant cell with bacterial infection.The length of these vector expression elements is different with characteristic.According to used host-vector integral body, can use many suitable any one that transcribe and translate in the element.

A plurality of levels that different hereditary signals and processing incident controlling gene are expressed (as, DNA transcribes and mRNA (mRNA) translation).

DNA transcribes the existence that depends on promotor, and the latter is a kind of dna sequence dna, and it is synthetic that therefore the combination of guide RNA polymerase also starts mRNA.The dna sequence dna of eukaryotic promoter is different from the dna sequence dna of prokaryotic promoter.In addition, eukaryotic promoter and the hereditary signal of following can not be identified in protokaryon integral body or not play a role, and in addition, prokaryotic promoter is not identified in eukaryotic and does not play a role.

Similarly, the translation of mRNA in prokaryotes depends on the suitable existence that is different from Eukaryotic prokaryotes signal.MRNA effectively translates the ribosome bind site that needs to be called on the mRNA Shine-Dalgarno (" SD ") sequence in prokaryotes.This sequence is the mRNA nucleotide sequence that is positioned at initiation codon weak point before, and initiation codon often is AUG, the aminoterminal methionine of encoding proteins.The SD sequence also may combine with ribosomal to allow the ribosomal location of correction to start mRNA by forming double helix with rRNA with 3 of 16srRNA (rRNA) '-end is complementary.For making the maximized summary of gene expression, see Roberts and Lauer, Enzymology method, 68:473 (1979) is incorporated herein only for reference.

" intensity " of promotor (being that they start the ability of transcribing) is different.For reaching the purpose that clone gene is expressed, for obtain gene high-caliber transcribe and, expression therefore, it is necessary using strong promoter.According to used host cell integral body, can use any one in many suitable promotors.For example,, in the time of in its phage or the plasmid, can use following promotor when cloning in Escherichia coli, as the T7 phage promoter, lac promotor, trp promotor, recA promotor, rRNA promotor, the P of escherichia coli _RAnd P _LPromotor and other promotor include but not limited to, lacUV5, and ompF, bla, lpp etc. transcribe with the high level that instructs adjacent dna fragmentation.In addition, can use heterozygote trp-lacUV5 (tac) promotor for preparing by recombinant DNA or other synthetic DNA technology or other escherichia coli promoter to be provided for inserting gene transcription.

Can select bacterial host cell bacterial strain and the expression vector that suppresses promoter function if need not to induce especially.In some operation, adding special inducer is essential to inserting effectively transcribing of DNA.For example, the lac operon is induced by adding lactose or IPTG (isopropylthio-).Other operon of many kinds, as trp, pro, etc., under different control.

Special initial signal also is transcribing and translate being used for efficient gene of needing in prokaryotic.These are transcribed and " intensity " of translation initiation signal can be inequality, and this " intensity " reaches synthetic albumen by quantitate gene specificity mRNA and measured respectively.The DNA expression vector that contains promotor also can contain various " by force " transcribes and/or any combination of translation initiation signal.For example, in Escherichia coli effectively translation need initiation codon (ATG) 5 ' Shine-Dalgarno (SD) sequence of about 7-9 base so that ribosome bind site to be provided.Therefore, can utilize and to be made up by any SD-ATG that the host cell ribosome utilizes.Such combination includes but not limited to from the cro gene of escherichia coli or N gene, or makes up from the SD-ATG of Escherichia coli tryptophan (tryptophan) E.D.C.B or A gene.In addition, can use by recombinant DNA or comprise any SD-ATG combination that other technology of mixing synthesizing ribonucleotide is prepared.

Express integral body in case the isolated DNA molecule of coding hypersensitive response elicitor polypeptide or albumen is cloned into, it just easily is incorporated into host cell.Mixing like this can be finished by above-mentioned various forms of being converted according to carrier/host cell integral body.The suitable hosts carrier cell includes, but are not limited to bacterium, virus, yeast, mammalian cell, insect, plant etc.

The inventive method can be used for handling various plants to give pathogen-resistance.Suitable plant comprises dicotyledonous and monocotyledon.More in particular, useful crops plant can comprise: paddy rice, wheat, barley, rye, cotton, sunflower, peanut, corn, potato, sweet potato, broad bean, pea, witloof, lettuce, corn sow thistle, cabbage, cauliflower, brocoli, radish, radish, spinach, onion, garlic, eggplant, pepper, celery, carrot, custard squash, pumpkin, zucchini, cucumber, apple, pears, muskmelon, strawberry, grape, raspberry, pineapple, soya bean, tobacco, tomato, Chinese sorghum, and sugarcane.The example of suitable decorative plant has: mouseearcress (Arabidopsis thaliana), African purple lettuce tongue belong to (Saintpaulia), Petunia, Pelargonium, poinsettia genus, Chrysanthemum, Carnation and one hundred days Chrysanthemum.

The method of giving phytopathogen antibody according to the present invention is useful in giving the resistance that multiple pathogene is comprised virus, bacterium and fungi.

Can finish by method of the present invention the resistance of following virus especially: tobacco mosaic virus and tomato mosaic virus.

Also can give plant to the resistance of following bacterium especially: the mutation of causing a disease of eggplant pseudomonas, pseudomonas syringae, tabaci, and Xanthomonas campestris mutation pelargonii according to the present invention.

The method of the application of the invention can make plant especially following fungi be had resistance: fusarium oxysporum and phytophthora infestan.

Method of the present invention can be finished by using hypersensitive response elicitor polypeptide or albumen to the whole bag of tricks of all or part of pending plant.This can (but needn't) comprises invades the hypersensitive response elicitor polypeptide or albumen enters plant.Suitable using method comprises that high pressure or low pressure spray, injection and the contiguous blade injury of inducer when using that carry out.Those skilled in the art can dream up other suitable using method, as long as it can make hypersensitive response elicitor polypeptide or albumen contact effectively with plant cell.

Hypersensitive response elicitor polypeptide or albumen can be applied to plant separately or with the mixture with other material according to the present invention.

One aspect of the present invention comprises and is used for giving the containing at the hypersensitive response elicitor polypeptide of carrier or the composition of albumen of plant pathogen resistance.Suitable carriers comprises the water or the aqueous solution.In this embodiment, composition contains greater than 500nM hypersensitive response elicitor polypeptide or albumen.

Although be not essential, said composition can contain extra additive and comprise fertilizer, insecticide, fungicide and composition thereof.Suitable fertilizer comprises (NH ₄) ₂NO ₃The example of suitable insecticide is the malathion.Useful fungicide comprises captan.

Other suitable additive comprises buffer, humidizer, reaches rubbing agent.Can use these materials with auxiliary method of the present invention.

The resistance of the anti-southern bacterialo wilt disease (aeruginosa eggplant) of the tomato that EXAMPLE Example 1-Harpin-induces

Plant in the greenhouse and handle below tomato seedling in all ages work of 2 on 8 * 15cm level land: 20 strain plants are used for a kind of of following six kinds of processing, and this handles called after A to F, and is described below:

(A) the thick harpin of 200 μ g/ml (being hypersensitive response elicitor polypeptide or the albumen) goods (Z-M.Wei of about 100 μ l, " Harpin; separate the hypersensitive response elicitor of starch Erwinia preparation by phytopathogen ", science 257:85-88 (1992) is incorporated herein only for reference) immerse in the minimum true leaf of every strain seedling.

(B) harpin goods identical with being used for (A) spray to the blade surface of seedling with the diamond dust of 400-mesh and rub gently with thumb then.

(C) bacillus coli DH 5 (pCPP 430) (collection of illustrative plates of plasmid vector pCPP 430 is seen Fig. 1) is incubated at the LB medium to OD ₂₆₀=0.7.Centrifugal culture and being resuspended in then in the kaliumphosphate buffer of 5mM of pH 6.5.The cell suspension of about 100 μ l is immersed in each blade of seedling.

(D) with (C) in the same bacillus coli DH 5 (pCPP 430 ∷ hrpN) (collection of illustrative plates of plasmid vector pCPP 430 ∷ hrpN is seen Fig. 1) that uses.Cultured cell, and used suspension and inoculum concentration and (C) middle describe identical.

(E) for bacillus coli DH 5 (pCPP9) (see figure 2), cultured cell and used suspension and inoculum concentration are with identical described in (C).

(F) blade is immersed with the 5mM kaliumphosphate buffer as (C) described in.

With the pathogenetic bacteria of attacking, aeruginosa eggplant bacterial strain K60 is incubated at KingShi medium B to OD ₂₆₀=0.7 (about 10 ⁸Cfu/ml).Centrifugal culture also is resuspended in the kaliumphosphate buffer of 100 volume 5mM to final concentration about 1 * 10 ⁶Cfu/ml.

, it is extracted and downcut about 1 centimetre root with harpin or bacterial treatment after three days tomato children bacterium with scissors.Then seedling was immersed among the aeruginosa eggplant K60 3 minutes.The plant that inoculated is planted again in identical canister.Plant is placed the greenhouse, and at postvaccinal 7 days record disease incidents.A. the influence of handling with harpin

After 24 hours, only those blade-section atrophys of immersing with harpin or bacillus coli DH 5 (pCPP 430).The blade that sprays with harpin and diamond dust only demonstrates mottled necrosis.B. handle the withered influence that forms of southern bacterium with harpin.

There is not a strain to show any symptom (table 1) after one week in the plant of 20 strains immersion harpin at inoculation aeruginosa eggplant K60.Strain plant in 20 strains demonstrates the dwarfing symptom.Yet, have 7 strains to show the dwarfing symptom in the plant of 20 strains immersion buffer solution.(pCPP 430 with bacillus coli DH 5 with comparing with the plant of buffer solution processing ^-) transposons-induced mutation of induced hypersensitivity atrophy (a kind of can not) or bacillus coli DH 5 (pCPP9) handle and do not demonstrate significant difference.The bacillus coli DH 5 (pCPP 430) that these results show harpin or produce harpin is induced the southern bacillary withered resistance that aeruginosa eggplant K60 is caused in the tomato plant.Table 1. is with the disease incident of aeruginosa eggplant K60 inoculation tomato seedling after 7 and 14 days.

The plant number

Process downgrading the healthy healthy A.Harpin of dwarfing in 7 days 14 days immerses 0 20 2 18B.Harpin and sprays withered 11 pathogen-free domestic 0 20 0 20 of withered 13F. buffer solution 7 13 8+1 of 1 19 3 17C. bacillus coli DH 5s (pCPP430), 2 18 3 17D. bacillus coli DH 5s (pCPP430), 4 16 7 13E. bacillus coli DH 5s (pCPP9), 5 15 6+1

In 4 weeks of inoculation back, the plant of handling with harpin or bacillus coli DH 5 (pCPP 430) is than the plant height and width of handling with buffer solution.Average height by 10 strain plants of harpin or buffer solution immersion is listed in table 2.After table 2. is handled with harpin or buffer solution, red with west, aeruginosa eggplant K60 inoculation 4 week back

The height of persimmon plant (cm).

Immerse buffer solution and immerse Harpin immersion buffer solution

Do not inoculate with the K60 inoculation and inoculate with K60

36 32 11

41 29 21

35 38 33

34 35 12

39 37 15

35 33 32

36 22 25

35 35 15

41 40 37

The tomato that 37 29 38 average 36.9 33 23.9 embodiment 2-Harp-induce is to the resistance of southern bacterialo wilt disease aeruginosa eggplant

Except aeruginosa eggplant K60 concentration is about 5 * 4 ⁴Beyond the cfu/ml, be used for immersing and all methods of inoculating all with embodiment 1 describe identical.

After 15 days, immersing has the plant of buffer solution to show symptom with aeruginosa eggplant K60 inoculation.Six strains in the 20 strain plants show short and small symptom after 15 days; 2 strain plants are withered after 21 days.Withered plant is finally dead.Yet neither one shows the dwarfing symptom in the plant that 20 strain harpin handle.After inoculating for 3 weeks, 3 strains show the dwarfing symptom in the plant that 20 strain harpin handle.After three weeks, it is possible that plant may be lost its induction of resistance.With the same in first experiment, whole girth (girth) of harpin processing plant and height are greater than the plant of handling with buffer solution.The tomato that embodiment 3-Harpin-induces is anti-to southern bacterialo wilt disease eggplant pseudomonas

The property

Except add extra eggplant pseudomonas K60 inoculum in the canister of the tomato plant that contains processing, this experiment is similar to Example 1.

Harpin is immersed in the tomato seedling in 2 ages in week.In two groups of plants every group immerses the harpin of about 200 μ l is suspended in the concentration of about 200 μ g/ml in the solution of kaliumphosphate buffer of 5mM.Immerse 20 strain tomato seedling altogether, the tomato seedling of similar number is immersed with buffer solution.Two days later, by the root immersion plant is inoculated with eggplant pseudomonas K60.The plant of harpin-or buffer solution-immersion extracted from soil mixture and cut off a spot of and then with in the remaining suspension that immerses eggplant pseudomonas K60 3 minutes with scissors.The concentration of bacterial cell suspension about 5 * 10 ⁸Cfu/ml.Seedling is planted again in identical canister.The bacterial suspension of extra 3ml is joined in the soil of each 4-inch diameter canister.Counting disease incident after one week.All experiments are all carried out in having the temperature controlled greenhouse of restriction.

After 3 weeks, the plant recovery that 11 strains in the tomato plant that 20 strain buffer solutions immerse are dead and 2 strains are withered is downgraded but keep serious.To compare only 4 strain plant growings normal with the tomato that does not have inoculation.Yet 15 strains are seemingly healthy in the plant that harpin handles; Inoculation back strain 3 Wednesdays dwarfing plants and 2 strain plants are withered.These the results are summarized in the following table 3.The resistance of the bacterialo wilt disease that the table 3.Harpin tomato of inducing causes the eggplant pseudomonas

Postvaccinal all numbers are processed 12 3Harpin healthy 20 17 15 withered 012 and are downgraded 023 buffer solutions healthy 854 withered 8 12 13 and downgrade tobacco that 433 embodiment 4-Harpin induce to the resistance of tobacco mosaic virus (TMV)

It is separating among the starch Erwinia harpin of 200 μ g/ml that the low level leaf of one group of 4 week tobacco seedling in age (cultivar, Xanthi have the N gene) immerses concentration.After three days, attack plant with tobacco mosaic virus (" TMV ").Use the virus (5 μ g and 100 μ g/ml) of 2 kinds of concentration.The viral suspension of about 50 μ l places on the upper tobacco smoked sheet.This blade spreads with the 400-purpose diamond dust and the blade that rubs lightly.Every kind of concentration is tested on three strain plants.Damage and report three means (table 4) on the blade in back 4 days of inoculation and 2 days counting gangrenosum acnes subsequently.Be difficult to distinguish single damage at the 10th day because some gangrenosum acne damage is fused to together.As if the damage number that therefore, records be less than the number that recorded at the 7th day.The blade that the scope of gangrenosum acne damage is handled greater than harpin in the blade of handling with buffer solution.The tobacco induced of table 4.Harpin is to from the resistance with the TMV of 5 μ g/ml virus inoculations

The mean of damage/blade is handled 4 days 7 days 10 days Harpin 21 32 35 buffer solutions 67 102 76

When the tobacco mosaic virus inoculum density is 100 μ g/ml, no significant difference between the local damage number that forms in the tobacco that harpin handles and buffer solution is handled.The tomato that embodiment 5-Harpin induces is to the resistance of Fusarium fusarium wilt

The tomato plant in six ages in week presses that embodiment 3 is described to be handled with harpin.Fungal pathogens, fusarium oxysporum is incubated in the lima bean agar medium 5 days in 27 ℃.Two complete agar plates with mycelia mixed 2 minutes in the kaliumphosphate buffer of 20ml 5mM.The tomato plant roots that harpin or buffer solution are handled is damaged to canister soil by inserting a timber.Then, the fungi suspension of 3ml is poured in 4 inches the soil of every canister.The plant of inoculation remains in 24 ℃ of greenhouses that control environment and illumination every day 16 hours.Inoculate and write down disease incident after 7 days.Every kind of processing is applied to 10 strain plants.The results are shown in following table 5.Table 5. harpin or buffer solution are handled the influence to tomato Fusarium fusarium wilt

Shown in inoculation after the timetable plant number (in 10 strains) that reveals withered symptom handle tobacco that 7 days 10 days 15 days 20 days Harpin 1244 (dead 1) buffer solution 3677 (dead 4) embodiment 6-Harpin induce to wildfire (pseudomonas syringae cause a disease mutation tabaci)

Resistance

Harpin is immersed in single group low level leaf of 4 week tobacco plants in age (20cm height).After 3 days, the suspension of the pathogenic mutation tabaci of pseudomonas syringae is immersed in single group superior leaf.After 4 days, the record disease incident is as listed in the table 6.After handling with harpin, table 6. low level leaf inoculates with the pathogenic mutation tabaci of pseudomonas syringae

The infection symptoms p.s.tabaci concentration of wildfire is not handled with Harpin and is handled 10 with Harpin in the tobacco leaf ⁴Asymptomatic necrosis of cfu/m and saturated with water 10 ⁵Asymptomatic necrosis of cfu/ml and saturated with water 10 ⁶Asymptomatic necrosis of cfu/ml and saturated with water 10 ⁷Asymptomatic necrosis of cfu/ml and saturated with water 10 ⁸The Mang ox seedling that cfu/ml is downright bad downright bad and saturated with water embodiment 7-Harpin-induces belongs to (Pelargonium hortorum) to carefully

The resistance of bacterium property tikka (Xanthomonas campestris cause a disease mutation pelargonii)

This experiment goes root Mang ox seedling to belong to carry out in the artificial soil mixture in single 4 " or 6 " canister to be incubated in the greenhouse.2 low level leaves on every strain plant are with the kaliumphosphate buffer pH 6.5 (contrast) of 0.05M, or harpin or bacillus coli DH 5 (pCPP 430) (complete clone separate starch Erwinia gene cluster) suspension immerses.After the immersion 2～7 days, all plants are with the bacterial leaf spot substance, the cause a disease pure culture inoculation of mutation pelargonii of Xanthomonas campestris.Bacterial suspension (5 * 10 ⁶Cfu/ml) be sprayed to upper and the next leaf surface of plant with low pressure.Every kind of processing is applied to 2 groups of plants (called after in table 7 " A " and " B ").Plant was maintained in the confined chamber 48 hours and replenished spraying by cold spray painting day with fog.Then, before analysis of disease takes place, plant is maintained at the little stool in greenhouse last 10 day, control on every side humidity and 23 ℃～32 ℃ temperature.Table 7.harpin conciliates starch Erwinia hrp gene cluster Mang ox seedling is belonged to bacillary tikka

The influence that forms.

Handle and handled 7 days 5 days 4 days 3 days 2 days with the time that Xanthomonas campestris causes a disease between the mutation pelargonii inoculation

Plant plant plant plant plant

A B A B A B A B A B buffer solution 3 ^*55432434 5Harpin, 000000100 0DH5 (pCPP430), 00 NT NT 000110 ^*Numeral in the table showed the number of disease symptoms (significant downright bad, chlorisis or withered) blade in back 10 days for inoculation.Several harpins of embodiment 8-are inducing the open country that the pathogenic mutation tabaci of pseudomonas syringae is caused

Activity in the sick resistance of fire

Tobacco plant (Nicotiana tabacum var.Xanthi) is planted in the greenhouse.When 4 ages in week, the harpin goods are immersed in two low level blades of single group of every strain plant.12 strain plants are handled with every kind of harpin goods, and three strains are to be used to prepare the identical kaliumphosphate buffer processing of harpins.In the group that immerses blade with the harpin goods, form hypersensitive necrosis in 24 hours, but do not form hypersensitive necrosis in the group with the buffer solution processing.

After harpin handles 7,10,11 and 12 days, all plants inoculate 10 by the superior leaf that immerses in every group ⁴-10 ⁶The pseudomonas syringae of the cell/ml mutation tabaci suspension that causes a disease.Plant was cultivated in the greenhouse 7 days before assess disease forms.The results are shown in following table 8:

Fate between table 8Harpin source processing and the inoculation

12 11 10 7log[inoculation] 456456456456 nothings (buffer solution)++ ++++ ++++ ++++ ++ pseudomonas syringae--+--+--+--+chrysanthemum Erwinia--+--+--+--+solution starch Erwinia--+-----+--+

-=do not have symptom,

+=downright bad and have yellow dizzy, typical wildfire

++=serious downright bad and have yellow dizzy, typical wildfire

The result shows that the harpin goods from three kinds of bacteriums all are effective in inducing the resistance of wildfire substance.The plant of handling with any harpin does not show any symptom after using two kinds low inoculum densities.After higher concentration inoculation, even more serious than symptom in the plant of harpin processing with symptom in the plant of buffer solution processing.The resistance of the late blight that phytophthora infestan is caused that embodiment 9-Harpin induces.

The late blight pathogene mainly influences potato and tomato.It causes notorious Irish potato famine.Harpin is inducing the tomato seedling that the usefulness of the activity in this pathogen-resistance is planted in the greenhouse to be measured.The seedling (cultivar " Mama Mia ", about 6～8 inches high) in three ages in week is handled with harpin and inoculates with phytophthora infestan subsequently.Two groups of low level blades of every strain plant immerse with bacillus coli DH 5 (pCPP 430) suspension or the kaliumphosphate buffer of harpin solution, generation and secretion harpin.

Immerse after 2,3 or 4 days, plant is with the mycelium suspension inoculation of phytophthora infestan.Use is to No. 7, poisonous bacterial strain U.S. of tomato height.The mycelium suspension prepares by be mixed in 21 ℃ of fungies of cultivating 2 days gently on agar plate of 2 barleys.Blade upper surface and the lower surface of the plant of this suspension brush to every strain being handled with artist's thick paintbrush.

The plant of handling and inoculating is designed to keep the having in the mist cell of special structure of 20-23 ℃ of temperature and cultivates in the greenhouse, keep high relative moisture simultaneously.Humidity by several on pure water the cold spray painting day with fog with maximum speed operation provide.Calculate disease incidence rate with the phytophthora infestan inoculation after 13 days, and will the results are shown in following table 9.Every kind of processing is applied to the single plant of 4 strains.Table 9. inoculation is present in the number of late blight damage in the tomato blade after 13 days.Fate between processing and the inoculation

4 3 2

Plant A B C D A B C D A B C D buffer solution 32001220004 1Harpin, 00100001210 0DH5 (pCPP430) 000102210110

Processing with harpin has reduced the number that forms damage in the plant in all intervals of handling and inoculating.To produce and to secrete DH5 (pCPP 430) processing of harpin, the late blight damage number of formation also reduces by in advance.

Although describe the present invention for illustrative purposes in detail, should be understood that this details only is for illustrative purposes, and those skilled in the art can make change under the situation that does not depart from the spirit and scope of the present invention that limited by following claim.

Sequence table (1) general information:

(i) applicant: Connell research foundation, company

(ii) invention exercise question: the resistance that allergy is induced in plant

(iii) sequence number: 9

(iv) address:

(A) address: Nixon, Hargrave, Devans ﹠amp; Doyle LLP

(B) street: Clinton Square, P.O.Box 1051

(C) city: Rochester

(D) state: New York

(E) country: the U.S.

(F) postcode: 14603

(v) computer-reader form:

(A) media type: floppy disk

(B) calculator: IBM PC compatible

(C) operating system: PC-DOS/MS-DOS

(D) software: patent discharges #1.0, #1.3 version

(vi) current request for data:

(A) application number:

(B) submission date:

(C) classification:

(vii) priority requisition data:

(A) application number: the U.S. 08/475,775

(B) submission date: June 7 nineteen ninety-five

(viii) lawyer/proxy information:

(A) name: Goldman, Michael L.

(B) number of registration: 30,727

(C) reference/catalog number (Cat.No.) of trying a case fully: 19603/10051

(ix) telecom information:

(A) phone: (716) 263-1304

(B) fax: (716) 263-1600 (2) Sequence Identification number 1 information:

(i) sequence signature:

(A) length: 338 amino acid

(B) type: amino acid

(C) chain:

(D) topology: linearity

(ii) molecule type: albumen

(xi) sequence description: Sequence Identification 1:Met Gln Ile Thr Ile Lys Ala His Ile Gly Gly Asp Leu Gly Val Ser1 5 10 15Gly Leu Gly Ala Gln Gly Leu Lys Gly Leu Asn Ser Ala Ala Ser Ser

20 25 30Leu?Gly?Ser?Ser?Val?Asp?Lys?Leu?Ser?Ser?Thr?Ile?Asp?Lys?Leu?Thr

35 40 45Ser?Ala?Leu?Thr?Ser?Met?Met?Phe?Gly?Gly?Ala?Leu?Ala?Gln?Gly?Leu

85 90 95Ser?Gly?Gly?Asp?Ala?Leu?Ser?Lys?Met?Phe?Asp?Lys?Ala?Leu?Asp?Asp

100 105 110Leu?Leu?Gly?His?Asp?Thr?Val?Thr?Lys?Leu?Thr?Asn?Gln?Ser?Asn?Gln

115 120 125Leu?Ala?Asn?Ser?Met?Leu?Asn?Ala?Ser?Gln?Met?Thr?Gln?Gly?Asn?Met

165 170 175Ala?Gly?Gly?Leu?Gln?Gly?Leu?Ser?Gly?Ala?Gly?Ala?Phe?Asn?Gln?Leu

180 185 190Gly?Asn?Ala?Ile?Gly?Met?Gly?Val?Gly?Gln?Asn?Ala?Ala?Leu?Ser?Ala

195 200 205Leu?Ser?Asn?Val?Ser?Thr?His?Val?Asp?Gly?Asn?Asn?Arg?His?Phe?Val

245 250 255Ser?Ser?Pro?Lys?Thr?Asp?Asp?Lys?Ser?Trp?Ala?Lys?Ala?Leu?Ser?Lys

260 265 270Pro?Asp?Asp?Asp?Gly?Met?Thr?Gly?Ala?Ser?Met?Asp?Lys?Phe?Arg?Gln

275 280 285Ala?Met?Gly?Met?Ile?Lys?Ser?Ala?Val?Ala?Gly?Asp?Thr?Gly?Asn?Thr

325 330 335Asn Ala (2) Sequence Identification number 2 information:

(i) sequence signature:

(A) length: 2141 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topology: linearity

(ii) molecule type: DNA (genome)

( xi ) ：2：CGATTTTACC CGGGTGAACG TGCTATGACC GACAGCATCA CGGTATTCGA CACCGTTACG 60GCGTTTATGG CCGCGATGAA CCGGCATCAG GCGGCGCGCT GGTCGCCGCA ATCCGGCGTC 120GATCTGGTAT TTCAGTTTGG GGACACCGGG CGTGAACTCA TGATGCAGAT TCAGCCGGGG 180CAGCAATATC CCGGCATGTT GCGCACGCTG CTCGCTCGTC GTTATCAGCA GGCGGCAGAG 240TGCGATGGCT GCCATCTGTG CCTGAACGGC AGCGATGTAT TGATCCTCTG GTGGCCGCTG 300CCGTCGGATC CCGGCAGTTA TCCGCAGGTG ATCGAACGTT TGTTTGAACT GGCGGGAATG 360ACGTTGCCGT CGCTATCCAT AGCACCGACG GCGCGTCCGC AGACAGGGAA CGGACGCGCC 420CGATCATTAA GATAAAGGCG GCTTTTTTTA TTGCAAAACG GTAACGGTGA GGAACCGTTT 480CACCGTCGGC GTCACTCAGT AACAAGTATC CATCATGATG CCTACATCGG GATCGGCGTG 540GGCATCCGTT GCAGATACTT TTGCGAACAC CTGACATGAA TGAGGAAACG AAATTATGCA 600AATTACGATC AAAGCGCACA TCGGCGGTGA TTTGGGCGTC TCCGGTCTGG GGCTGGGTGC 660TCAGGGACTG AAAGGACTGA ATTCCGCGGC TTCATCGCTG GGTTCCAGCG TGGATAAACT 720GAGCAGCACC ATCGATAAGT TGACCTCCGC GCTGACTTCG ATGATGTTTG GCGGCGCGCT 780GGCGCAGGGG CTGGGCGCCA GCTCGAAGGG GCTGGGGATG AGCAATCAAC TGGGCCAGTC 840TTTCGGCAAT GGCGCGCAGG GTGCGAGCAA CCTGCTATCC GTACCGAAAT CCGGCGGCGA 900TGCGTTGTCA AAAATGTTTG ATAAAGCGCT GGACGATCTG CTGGGTCATG ACACCGTGAC 960CAAGCTGACT AACCAGAGCA ACCAACTGGC TAATTCAATG CTGAACGCCA GCCAGATGAC 1020CCAGGGTAAT ATGAATGCGT TCGGCAGCGG TGTGAACAAC GCACTGTCGT CCATTCTCGG 1080CAACGGTCTC GGCCAGTCGA TGAGTGGCTT CTCTCAGCCT TCTCTGGGGG CAGGCGGCTT 1140GCAGGGCCTG AGCGGCGCGG GTGCATTCAA CCAGTTGGGT AATGCCATCG GCATGGGCGT 1200GGGGCAGAAT GCTGCGCTGA GTGCGTTGAG TAACGTCAGC ACCCACGTAG ACGGTAACAA 1260CCGCCACTTT GTAGATAAAG AAGATCGCGG CATGGCGAAA GAGATCGGCC AGTTTATGGA 1320TCAGTATCCG GAAATATTCG GTAAACCGGA ATACCAGAAA GATGGCTGGA GTTCGCCGAA 1380GACGGACGAC AAATCCTGGG CTAAAGCGCT GAGTAAACCG GATGATGACG GTATGACCGG 1440CGCCAGCATG GACAAATTCC GTCAGGCGAT GGGTATGATC AAAAGCGCGG TGGCGGGTGA 1500TACCGGCAAT ACCAACCTGA ACCTGCGTGG CGCGGGCGGT GCATCGCTGG GTATCGATGC 1560GGCTGTCGTC GGCGATAAAA TAGCCAACAT GTCGCTGGGT AAGCTGGCCA ACGCCTGATA 1620ATCTGTGCTG GCCTGATAAA GCGGAAACGA AAAAAGAGAC GGGGAAGCCT GTCTCTTTTC 1680TTATTATGCG GTTTATGCGG TTACCTGGAC CGGTTAATCA TCGTCATCGA TCTGGTACAA 1740ACGCACATTT TCCCGTTCAT TCGCGTCGTT ACGCGCCACA ATCGCGATGG CATCTTCCTC 1800GTCGCTCAGA TTGCGCGGCT GATGGGGAAC GCCGGGTGGA ATATAGAGAA ACTCGCCGGC 1860CAGATGGAGA CACGTCTGCG ATAAATCTGT GCCGTAACGT GTTTCTATCC GCCCCTTTAG 1920CAGATAGATT GCGGTTTCGT AATCAACATG GTAATGCGGT TCCGCCTGTG CGCCGGCCGG 1980GATCACCACA ATATTCATAG AAAGCTGTCT TGCACCTACC GTATCGCGGG AGATACCGAC 2040AAAATAGGGC AGTTTTTGCG TGGTATCCGT GGGGTGTTCC GGCCTGACAA TCTTGAGTTG 2100GTTCGTCATC ATCTTTCTCC ATCTGGGCGA CCTGATCGGT T 2141 ( 2 ) 3：

(i) sequence signature:

(A) length: 385 amino acid

(B) type: amino acid

(C) chain:

(D) topology: linearity

(ii) molecule type: albumen (xi) sequence description: Sequence Identification 3:Met Ser Leu Asn Thr Ser Gly Leu Gly Ala Ser Thr Met Gln Ile Ser1 5 10 15Ile Gly Gly Ala Gly Gly Asn Asn Gly Leu Leu Gly Thr Ser Arg Gln

20 25 30Asn?Ala?Gly?Leu?Gly?Gly?Asn?Ser?Ala?Leu?Gly?Leu?Gly?Gly?Gly?Asn

35 40 45Gln?Asn?Asp?Thr?Val?Asn?Gln?Leu?Ala?Gly?Leu?Leu?Thr?Gly?Met?Met

85 90 95Gly?Leu?Ser?Asn?Ala?Leu?Asn?Asp?Met?Leu?Gly?Gly?Ser?Leu?Asn?Thr

100 105 110Leu?Gly?Ser?Lys?Gly?Gly?Asn?Asn?Thr?Thr?Ser?Thr?Thr?Asn?Ser?Pro

115 120 125Leu?Asp?Gln?Ala?Leu?Gly?Ile?Asn?Ser?Thr?Ser?Gln?Asn?Asp?Asp?Ser

165 170 175Gln?Asp?Gly?Thr?Gln?Gly?Ser?Ser?Ser?Gly?Gly?Lys?Gln?Pro?Thr?Glu

180 185 190Gly?Glu?Gln?Asn?Ala?Tyr?Lys?Lys?Gly?Val?Thr?Asp?Ala?Leu?Ser?Gly

195 200 205Leu?Met?Gly?Asn?Gly?Leu?Ser?Gln?Leu?Leu?Gly?Asn?Gly?Gly?Leu?Gly

245 250 255Leu?Gly?Asn?Ala?Val?Gly?Thr?Gly?Ile?Gly?Met?Lys?Ala?Gly?Ile?Gln

260 265 270Ala?Leu?Asn?Asp?Ile?Gly?Thr?His?Arg?His?Ser?Ser?Thr?Arg?Ser?Phe

275 280 285Val?Asn?Lys?Gly?Asp?Arg?Ala?Met?Ala?Lys?Glu?Ile?Gly?Gln?Phe?Met

325 330 335Lys?Pro?Asp?Asp?Asp?Gly?Met?Thr?Pro?Ala?Ser?Met?Glu?Gln?Phe?Asn

340 345 350Lys?Ala?Lys?Gly?Met?Ile?Lys?Arg?Pro?Met?Ala?Gly?Asp?Thr?Gly?Asn

355 360 365Gly?Asn?Leu?Gln?His?Ala?Val?Pro?Val?Val?Leu?Arg?Trp?Val?Leu?Met

370 375 380Pro385 (2) Sequence Identification number 4 information:

(i) sequence signature:

(A) length: 1158 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topology: linearity

(ii) molecule type: DNA (genome)

( xi ) ：4：ATGAGTCTGA ATACAAGTGG GCTGGGAGCG TCAACGATGC AAATTTCTAT CGGCGGTGCG 60GGCGGAAATA ACGGGTTGCT GGGTACCAGT CGCCAGAATG CTGGGTTGGG TGGCAATTCT 120GCACTGGGGC TGGGCGGCGG TAATCAAAAT GATACCGTCA ATCAGCTGGC TGGCTTACTC 180ACCGGCATGA TGATGATGAT GAGCATGATG GGCGGTGGTG GGCTGATGGG CGGTGGCTTA 240GGCGGTGGCT TAGGTAATGG CTTGGGTGGC TCAGGTGGCC TGGGCGAAGG ACTGTCGAAC 300GCGCTGAACG ATATGTTAGG CGGTTCGCTG AACACGCTGG GCTCGAAAGG CGGCAACAAT 360ACCACTTCAA CAACAAATTC CCCGCTGGAC CAGGCGCTGG GTATTAACTC AACGTCCCAA 420AACGACGATT CCACCTCCGG CACAGATTCC ACCTCAGACT CCAGCGACCC GATGCAGCAG 480CTGCTGAAGA TGTTCAGCGA GATAATGCAA AGCCTGTTTG GTGATGGGCA AGATGGCACC 540CAGGGCAGTT CCTCTGGGGG CAAGCAGCCG ACCGAAGGCG AGCAGAACGC CTATAAAAAA 600GGAGTCACTG ATGCGCTGTC GGGCCTGATG GGTAATGGTC TGAGCCAGCT CCTTGGCAAC 660GGGGGACTGG GAGGTGGTCA GGGCGGTAAT GCTGGCACGG GTCTTGACGG TTCGTCGCTG 720GGCGGCAAAG GGCTGCAAAA CCTGAGCGGG CCGGTGGACT ACCAGCAGTT AGGTAACGCC 780GTGGGTACCG GTATCGGTAT GAAAGCGGGC ATTCAGGCGC TGAATGATAT CGGTACGCAC 840AGGCACAGTT CAACCCGTTC TTTCGTCAAT AAAGGCGATC GGGCGATGGC GAAGGAAATC 900GGTCAGTTCA TGGACCAGTA TCCTGAGGTG TTTGGCAAGC CGCAGTACCA GAAAGGCCCG 960GGTCAGGAGG TGAAAACCGA TGACAAATCA TGGGCAAAAG CACTGAGCAA GCCAGATGAC 1020GACGGAATGA CACCAGCCAG TATGGAGCAG TTCAACAAAG CCAAGGGCAT GATCAAAAGG 1080CCCATGGCGG GTGATACCGG CAACGGCAAC CTGCAGCACG CGGTGCCGGT GGTTCTTCGC 1140TGGGTATTGA TGCCATGA 1158 ( 2 ) 5：

(i) sequence signature:

(A) length: 341 amino acid

(B) type: amino acid

(C) chain:

(D) topology: linearity

(ii) molecule type: albumen

(xi) sequence description: Sequence Identification 5:Met Gln Ser Leu Ser Leu Asn Ser Ser Ser Leu Gln Thr Pro Ala Met1 5 10 15Ala Leu Val Leu Val Arg Pro Glu Ala Glu Thr Thr Gly Ser Thr Ser

20 25 30Ser?Lys?Ala?Leu?Gln?Glu?Val?Val?Val?Lys?Leu?Ala?Glu?Glu?Leu?Met

35 40 45Arg?Asn?Gly?Gln?Leu?Asp?Asp?Ser?Ser?Pro?Leu?Gly?Lys?Leu?Leu?Ala

50 55 60Lys?Ser?Met?Ala?Ala?Asp?Gly?Lys?Ala?Gly?Gly?Gly?Ile?Glu?Asp?Val65 70 75 80Ile?Ala?Ala?Leu?Asp?Lys?Leu?Ile?His?Glu?Lys?Leu?Gly?Asp?Asn?Phe

85 90 95Gly?Ala?Ser?Ala?Asp?Ser?Ala?Ser?Gly?Thr?Gly?Gln?Gln?Asp?Leu?Met

100 105 110Thr?Gln?Val?Leu?Asn?Gly?Leu?Ala?Lys?Ser?Met?Leu?Asp?Asp?Leu?Leu

115 120 125Thr?Lys?Gln?Asp?Gly?Gly?Thr?Ser?Phe?Ser?Glu?Asp?Asp?Met?Pro?Met

165 170 175Leu?Asp?Gly?Asp?Glu?Thr?Ala?Ala?Phe?Arg?Ser?Ala?Leu?Asp?Ile?Ile

180 185 190Gly?Gln?Gln?Leu?Gly?Asn?Gln?Gln?Ser?Asp?Ala?Gly?Ser?Leu?Ala?Gly

195 200 205Thr?Gly?Gly?Gly?Leu?Gly?Thr?Pro?Ser?Ser?Phe?Ser?Asn?Asn?Ser?Ser

245 250 255Arg?Gly?Leu?Gln?Ser?Val?Leu?Ala?Gly?Gly?Gly?Leu?Gly?Thr?Pro?Val

260 265 270Asn?Thr?Pro?Gln?Thr?Gly?Thr?Ser?Ala?Asn?Gly?Gly?Gln?Ser?Ala?Gln

275 280 285Asp?Leu?Asp?Gln?Leu?Leu?Gly?Gly?Leu?Leu?Leu?Lys?Gly?Leu?Glu?Ala

325 330 335Asn?Gln?Ala?Ala?Ala

340 (2) Sequence Identification number 6 information:

(i) sequence signature:

(A) length: 1026 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topology: linearity

(ii) molecule type: DNA (genome)

( xi ) ：6：ATGCAGAGTC TCAGTCTTAA CAGCAGCTCG CTGCAAACCC CGGCAATGGC CCTTGTCCTG 60GTACGTCCTG AAGCCGAGAC GACTGGCAGT ACGTCGAGCA AGGCGCTTCA GGAAGTTGTC 120GTGAAGCTGG CCGAGGAACT GATGCGCAAT GGTCAACTCG ACGACAGCTC GCCATTGGGA 180AAACTGTTGG CCAAGTCGAT GGCCGCAGAT GGCAAGGCGG GCGGCGGTAT TGAGGATGTC 240ATCGCTGCGC TGGACAAGCT GATCCATGAA AAGCTCGGTG ACAACTTCGG CGCGTCTGCG 300GACAGCGCCT CGGGTACCGG ACAGCAGGAC CTGATGACTC AGGTGCTCAA TGGCCTGGCC 360AAGTCGATGC TCGATGATCT TCTGACCAAG CAGGATGGCG GGACAAGCTT CTCCGAAGAC 420GATATGCCGA TGCTGAACAA GATCGCGCAG TTCATGGATG ACAATCCCGC ACAGTTTCCC 480AAGCCGGACT CGGGCTCCTG GGTGAACGAA CTCAAGGAAG ACAACTTCCT TGATGGCGAC 540GAAACGGCTG CGTTCCGTTC GGCACTCGAC ATCATTGGCC AGCAACTGGG TAATCAGCAG 600AGTGACGCTG GCAGTCTGGC AGGGACGGGT GGAGGTCTGG GCACTCCGAG CAGTTTTTCC 660AACAACTCGT CCGTGATGGG TGATCCGCTG ATCGACGCCA ATACCGGTCC CGGTGACAGC 720GGCAATACCC GTGGTGAAGC GGGGCAACTG ATCGGCGAGC TTATCGACCG TGGCCTGC AA 780TCGGTATTGG CCGGTGGTGG ACTGGGCACA CCCGTAAACA CCCCGCAGAC CGGTACGTCG 840GCGAATGGCG GACAGTCCGC TCAGGATCTT GATCAGTTGC TGGGCGGCTT GCTGCTCAAG 900GGCCTGGAGG CAACGCTCAA GGATGCCGGG CAAACAGGCA CCGACGTGCA GTCGAGCGCT 960GCGCAAATCG CCACCTTGCT GGTCAGTACG CTGCTGCAAG GCACCCGCAA TCAGGCTGCA 1020GCCTGA 1026 ( 2 ) 7：

(i) sequence signature:

(A) length: 344 amino acid

(B) type: amino acid

(C) chain:

(D) topology: linearity

(ii) molecule type: albumen

(xi) sequence description: Sequence Identification 7:Met Ser Val Gly Asn Ile Gln Ser Pro Ser Asn Leu Pro Gly Leu Gln1 5 10 15Asn Leu Asn Leu Asn Thr Asn Thr Asn Ser Gln Gln Ser Gly Gln Ser

20 25 30Val?Gln?Asp?Leu?Ile?Lys?Gln?Val?Glu?Lys?Asp?Ile?Leu?Asn?Ile?Ile

35 40 45Ala?Ala?Leu?Val?Gln?Lys?Ala?Ala?Gln?Ser?Ala?Gly?Gly?Asn?Thr?Gly

85 90 95Ala?Asn?Lys?Thr?Gly?Asn?Val?Asp?Asp?Ala?Asn?Asn?Gln?Asp?Pro?Met

100 105 110Gln?Ala?Leu?Met?Gln?Leu?Leu?Glu?Asp?Leu?Val?Lys?Leu?Leu?Lys?Ala

115 120 125Ala?Leu?His?Met?Gln?Gln?Pro?Gly?Gly?Asn?Asp?Lys?Gly?Asn?Gly?Val

165 170 175Gly?Ala?Gly?Ala?Gly?Gly?Ala?Gly?Gly?Gly?Val?Gly?Gly?Ala?Gly?Gly

180 185 190Ala?Asp?Gly?Gly?Ser?Gly?Ala?Gly?Gly?Ala?Gly?Gly?Ala?Asn?Gly?Ala

195 200 205Asp?Gly?Gly?Asn?Gly?Val?Asn?Gly?Asn?Gln?Ala?Asn?Gly?Pro?Gln?Asn

245 250 255Ala?Leu?Val?Gln?Met?Met?Gln?Gln?Gly?Gly?Leu?Gly?Gly?Gly?Asn?Gln

260 265 270Ala?Gln?Gly?Gly?Ser?Lys?Gly?Ala?Gly?Asn?Ala?Ser?Pro?Ala?Ser?Gly

275 280 285Ala?Asn?Pro?Gly?Ala?Asn?Gln?Pro?Gly?Ser?Ala?Asp?Asp?Gln?Ser?Ser

325 330 335Gln?Ser?Thr?Ser?Thr?Gln?Pro?Met

340 (2) Sequence Identification number 8 information:

(i) sequence signature:

(A) length: 1035 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topology: linearity

(ii) molecule type: DNA (genome)

( xi ) ：8：ATGTCAGTCG GAAACATCCA GAGCCCGTCG AACCTCCCGG GTCTGCAGAA CCTGAACCTC 60AACACCAACA CCAACAGCCA GCAATCGGGC CAGTCCGTGC AAGACCTGAT CAAGCAGGTC 120GAGAAGGACA TCCTCAACAT CATCGCAGCC CTCGTGCAGA AGGCCGCACA GTCGGCGGGC 180GGCAACACCG GTAACACCGG CAACGCGCCG GCGAAGGACG GCAATGCCAA CGCGGGCGCC 240AACGACCCGA GCAAGAACGA CCCGAGCAAG AGCCAGGCTC CGCAGTCGGC CAACAAGACC 300GGCAACGTCG ACGACGCCAA CAACCAGGAT CCGATGCAAG CGCTGATGCA GCTGCTGGAA 360GACCTGGTGA AGCTGCTGAA GGCGGCCCTG CACATGCAGC AGCCCGGCGG CAATGACAAG 420GGCAACGGCG TGGGCGGTGC CAACGGCGCC AAGGGTGCCG GCGGCCAGGG CGGCCTGGCC 480GAAGCGCTGC AGGAGATCGA GCAGATCCTC GCCCAGCTCG GCGGCGGCGG TGCTGGCGCC 540GGCGGCGCGG GTGGCGGTGT CGGCGGTGCT GGTGGCGCGG ATGGCGGCTC CGGTGCGGGT 600GGCGCAGGCG GTGCGAACGG CGCCGACGGC GGCAATGGCG TGAACGGCAA CCAGGCGAAC 660GGCCCGCAGA ACGCAGGCGA TGTCAACGGT GCCAACGGCG CGGATGACGG CAGCGAAGAC 720CAGGGCGGCC TCACCGGCGT GCTGCAAAAG CTGATGAAGA TCCTGAACGC GCTGGTGCAG 780ATGATGCAGC AAGGCGGCCT CGGCGGCGGC AACCAGGCGC AGGGCGGCTC GAAGGGTGCC 840GGCAACGCCT CGCCGGCTTC CGGCGCGAAC CCGGGCGCGA ACCAGCCCGG TTCGGCGGAT 900GATCAATCGT CCGGCCAGAA CAATCTGCAA TCCCAGATCA TGGATGTGGT GAAGGAGGTC 960GTCCAGATCC TGCAGCAGAT GCTGGCGGCG CAGAACGGCG GCAGCCAGCA GTCCACCTCG 1020ACGCAGCCGA TGTAA 1035 ( 2 ) 9：

(i) sequence signature:

(A) length: 26 amino acid

(B) type: amino acid

(C) chain:

(D) topology: linearity

(ii) molecule type: peptide

(xi) sequence description: Sequence Identification 9:Thr Leu Ile Glu Leu Met Ile Val Val Ala Ile Ile Ala Ile Leu Ala1 5 10 15Ala Ile Ala Leu Pro Ala Tyr Gln Asp Tyr

20 25

Claims

1. method of giving plant pathogen resistance comprises:

Hypersensitive response elicitor polypeptide or albumen with non-infectious form under polypeptide or albumen and condition that plant cell contacts put on plant.

According to the hypersensitive response elicitor polypeptide that the process of claim 1 wherein or albumen corresponding to separating the starch Erwinia, the chrysanthemum Erwinia from being selected from, pseudomonas syringae, aeruginosa eggplant, Xanthomonas campestris, and composition thereof the inducer polypeptide or the albumen of pathogene.

3. according to the method for claim 2, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from the chrysanthemum Erwinia.

4. according to the method for claim 3, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification number 1.

5. according to the method for claim 3, the molecular weight that hypersensitive response elicitor polypeptide wherein or albumen have 34kDa.

6. according to the method for claim 2, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or the albumen of explaining the starch Erwinia by oneself.

7. according to the method for claim 6, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 3.

8. according to the method for claim 6, the molecular weight that hypersensitive response elicitor polypeptide wherein or albumen have 37kDa.

9. according to the method for claim 2, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from pseudomonas syringae.

10. according to the method for claim 9, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 5.

11. according to the method for claim 9, the molecular weight that hypersensitive response elicitor polypeptide wherein or albumen have 34-35kDa.

12. according to the method for claim 2, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from aeruginosa eggplant.

13. according to the method for claim 12, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 7.

14. according to the method for claim 2, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from Xanthomonas campestris.

15. according to the method for claim 14, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 9.

16. be selected from dicotyledonous and monocotyledon according to the plant that the process of claim 1 wherein.

17. according to the method for claim 16, plant wherein is selected from paddy rice, wheat, barley, rye, cotton, sunflower, peanut, corn, potato, sweet potato, broad bean, pea, witloof, lettuce, corn sow thistle, cabbage, cauliflower, brocoli, radish, radish, spinach, onion, garlic, eggplant, pepper, celery, carrot, custard squash, pumpkin, zucchini, cucumber, apple, pears, muskmelon, strawberry, grape, raspberry, pineapple, soya bean, tobacco, tomato, Chinese sorghum and sugarcane.

18. according to the method for claim 16, plant wherein is selected from mouseearcress (Arabidopsisthaliana), African purple lettuce tongue belongs to (Saintpaulia), Petunia, Pelargonium, poinsettia genus, Chrysanthemum, Carnation and one hundred days Chrysanthemum.

19. the pathogene according to the plant opposing that the process of claim 1 wherein is selected from virus, bacterium, fungi and combination thereof.

20. according to the process of claim 1 wherein that described application realizes by spraying, injection or the time scratch blade when contiguous described application takes place.

21. be applied to plant as the composition that further comprises carrier according to hypersensitive response elicitor polypeptide that the process of claim 1 wherein or albumen.

22. according to the method for claim 21, carrier wherein is selected from the water and the aqueous solution.

23. according to the method for claim 21, composition wherein contains hypersensitive response elicitor polypeptide or the albumen greater than 500nM.

24. according to the method for claim 21, composition wherein further contains the additive that is selected from fertilizer, insecticide, fungicide and composition thereof.

25. be according to hypersensitive response elicitor polypeptide or the albumen that the process of claim 1 wherein.With the form of separating.

26. according to the hypersensitive response elicitor polypeptide that the process of claim 1 wherein or albumen as not causing disease and using with the bacterium of the genetic transformation of coding hypersensitive response elicitor polypeptide or albumen.

27. method according to claim 1, wherein hypersensitive response elicitor polypeptide or albumen are used as bacterium, this bacterium causes disease in some floristics, but desire to carry out not cause disease in the described kind of using at those, and contain the gene of coding hypersensitive response elicitor polypeptide or albumen.

28. according to the process of claim 1 wherein that described application causes polypeptide or albumen to immerse in the plant.

29. the pathogen-resistance plant, its cell contacts with non-infectious hypersensitive response elicitor polypeptide or albumen.

30. pathogen-resistance plant according to claim 29, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to separating the starch Erwinia from being selected from, the chrysanthemum Erwinia, pseudomonas syringae, aeruginosa eggplant, Xanthomonas campestris, and composition thereof the hypersensitive response elicitor polypeptide or the albumen of pathogene.

31. according to the pathogen-resistance plant of claim 30, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from the chrysanthemum Erwinia.

32. according to the pathogen-resistance plant of claim 31, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification number 1.

33. according to the pathogen-resistance plant of claim 30, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or the albumen of explaining the starch Erwinia by oneself.

34. according to the pathogen-resistance plant of claim 33, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 3.

35. according to the pathogen-resistance plant of claim 30, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from pseudomonas syringae.

36. according to the pathogen-resistance plant of claim 35, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 5.

37. according to the pathogen-resistance plant of claim 30, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from aeruginosa eggplant.

38. according to the pathogen-resistance plant of claim 37, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 7.

39. according to the pathogen-resistance plant of claim 30, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from Xanthomonas campestris.

40. according to the pathogen-resistance plant of claim 39, hypersensitive response elicitor polypeptide wherein or albumen have the amino acid sequence corresponding to Sequence Identification numbers 9.

41. according to the pathogen-resistance plant of claim 29, plant wherein is selected from unifacial leaf and dicotyledon.

42. according to the pathogen-resistance plant of claim 41, plant wherein is selected from paddy rice, wheat, barley, rye, cotton, sunflower, peanut, potato, sweet potato, broad bean, pea, witloof, lettuce, corn sow thistle, cabbage, cauliflower, brocoli, radish, radish, spinach, onion, garlic, eggplant, pepper, celery, carrot, custard squash, pumpkin, zucchini, cucumber, apple, pears, muskmelon, strawberry, grape, raspberry, pineapple, soya bean, tobacco, tomato, Chinese sorghum and sugarcane.

43. according to the pathogen-resistance plant of claim 41, plant wherein is selected from mouseearcress (Arabidopsis thaliana), African purple lettuce tongue belongs to (Saintpaulia), Petunia, Pelargonium, poinsettia genus, Chrysanthemum, Carnation and one hundred days Chrysanthemum.

44. according to the pathogen-resistance plant of claim 30, plant wherein is selected from virus, bacterium, fungi and combination thereof to its pathogene with resistance.

45. according to claim 29 pathogen-resistance plant, hypersensitive response elicitor polypeptide wherein or albumen form for separating.

46. according to the pathogen-resistance plant of claim 29, plant cell wherein with do not cause disease and contact with the bacterium of the genetic transformation of coding hypersensitive response elicitor polypeptide or albumen.

47. according to the pathogen-resistance plant of claim 29, plant cell wherein contacts with bacterium, this bacterium does not cause disease in this plant, but causes disease really in other floristics, and contains the gene of coding hypersensitive response elicitor polypeptide or albumen.

48. according to the pathogen-resistance plant of claim 29, plant is wherein infiltrated with polypeptide or albumen.

49. a composition that is used to give plant pathogen resistance comprises:

Noninfective hypersensitive response elicitor polypeptide or albumen and

Carrier.

50. composition according to claim 49, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to separating the starch Erwinia from being selected from, the chrysanthemum Erwinia, pseudomonas syringae, aeruginosa eggplant, Xanthomonas campestris, with and composition thereof the hypersensitive response elicitor polypeptide or the albumen of pathogene.

51. according to the composition of claim 50, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from the chrysanthemum Erwinia.

52. according to the composition of claim 50, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or the albumen of explaining the starch Erwinia by oneself.

53. according to the composition of claim 50, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from pseudomonas syringae.

54. according to the composition of claim 50, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from aeruginosa eggplant.

55. according to the composition of claim 50, hypersensitive response elicitor polypeptide wherein or albumen are corresponding to hypersensitive response elicitor polypeptide or albumen from Xanthomonas campestris.

56. according to the composition of claim 49, carrier wherein is selected from the water and the aqueous solution.

57. according to the composition of claim 49, composition wherein contains hypersensitive response elicitor polypeptide or the albumen greater than 500nM.

58. according to the composition of claim 49, composition wherein further contains the additive that is selected from fertilizer, insecticide, fungicide and composition thereof.

59. according to the composition of claim 49, hypersensitive response elicitor polypeptide wherein or albumen form for separating.

60. according to the composition of claim 49, hypersensitive response elicitor polypeptide wherein or albumen are produced or can be produced by it by the bacterium in the composition, described bacterium does not cause disease and with the genetic transformation of coding hypersensitive response elicitor polypeptide or albumen.

61. according to the composition of claim 49, allergy polypeptide wherein or albumen are produced or can be produced thus by the bacterium that can cause disease in plant and contain the gene of coding hypersensitive response elicitor polypeptide or albumen.